Co-reporter:Rui Zhang;Qianqian Wu
RSC Advances (2011-Present) 2017 vol. 7(Issue 67) pp:42549-42558
Publication Date(Web):2017/08/29
DOI:10.1039/C7RA03805D
Widespread exposure to ultrafine carbon black (UFCB) and its ability to cross the lung-blood barrier have raised concerns regarding its safety. Importantly, UFCB can bind with proteins in the biological fluid after entering a biological environment and immediately form a protein corona. The protein corona could govern its further fate in the biological environment. In this work, we investigated the effects of UFCB to the structure and activity of catalase (CAT) to explore its biocompatibility with CAT. FW 200 (13 nm) and tween 80 (T80) were used as the UFCB model and the dispersant. Electron microscopy and dynamic light scattering (DLS) were used to characterize the surface properties and size distribution of UFCB. Steady-state fluorescence combined with synchronous fluorescence and 3D fluorescence show that UFCB bound with CAT and formed the protein corona, resulting an exposure of the internal amino acids (mainly tryptophan and tyrosine) and a decrease of the hydrophobicity of the amino acids. The fluorescence lifetime measurement combined with UV-visible spectra measurement indicated that UFCB quenched the fluorescence of CAT statically and changed the framework of CAT. Circular dichroism (CD) spectra analysis indicated the increase of α-helical content and the decrease of β-sheet structure in catalase, which in turn make the activity of CAT reduce as shown in the enzyme activity assay. The study demonstrated the negative effects of UFCB on proteins and stressed the urgency to conduct more investigation on the biosafety of its application.
Co-reporter:Yang Liu;Zhaozhen Cao;Wansong Zong
RSC Advances (2011-Present) 2017 vol. 7(Issue 40) pp:24781-24788
Publication Date(Web):2017/05/05
DOI:10.1039/C7RA02963B
In this study, the toxic mechanism and effects of perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHS), and perfluorobutane sulfonate (PFBS) were investigated via spectroscopy, molecular modeling, and calorimetry techniques. Results showed that all three perfluoroalkyl sulfonates (PFASs) bound to human serum albumin (HSA) mainly through electrostatic forces and hydrogen bonds. The backbone and secondary structure of HSA did not significantly change after exposure to PFASs. It may be proposed that the binding changed the local structure around the active site and affected the esterase activity of HSA. Compared with the control group, the inhibited esterase activity of HSA decreased to 28.6%, 43.2%, and 54.4% under the exposure of PFOS, PFHS, and PFBS at 1.3 × 10−4 mol L−1, respectively. The ITC result reflected that the binding ability increased after lengthening of the carbon chain, which also explained the decreased esterase activity with the increased lengthening of carbon chain. The fluorescence spectra also indicated that the influence on the microenvironment of HSA decreased with the shortening of the carbon chain. This study provided evidence regarding the interaction mechanism and toxicity of PFASs towards HSA in vitro.
Co-reporter:Jing Wang, Jinhu Wang, Wei Song, Xinping Yang, Wansong Zong and Rutao Liu
Physical Chemistry Chemical Physics 2016 vol. 18(Issue 5) pp:3536-3544
Publication Date(Web):11 Jan 2016
DOI:10.1039/C5CP06100H
Cadmium adversely affects the biological function of the liver. Transferrin might be involved in the detoxification system of cadmium. However, owing to the lack of investigation of the molecular mechanism of cadmium conjugating to transferrin, the role of transferrin in cadmium detoxification in the liver and how transferrin undergoes conformational and functional changes upon cadmium binding are not clear. In this article, we demonstrated the potential role of transferrin in the protection of the mouse primary hepatocytes against cadmium toxicity. After the incubation of hepatocytes with 10 and 100 μM CdCl2, pretreatment with transferrin significantly attenuated the reduction of cell viability in comparison with the samples treated with CdCl2 alone. Furthermore, a detailed molecular mechanism investigation of the interaction of CdCl2 with transferrin was reported using biophysical methods. Multi-spectroscopic measurements showed that CdCl2 formed complexes with transferrin and caused structural and conformational changes of transferrin. Isothermal titration calorimetry measurements revealed that transferrin has two classes of binding sites with different binding constants for CdCl2 binding. Hydrophobic forces and electrostatic forces are the major driving forces of the interaction. Preferred specific binding sites on transferrin were identified by dialysis experiments, molecular docking studies and molecular dynamics simulations. Upon low CdCl2 concentration exposure, no content of iron was released from transferrin because CdCl2 preferentially binds to the surface of transferrin molecules. Upon higher CdCl2 concentration exposure, the release of iron content from transferrin was observed due to the interaction of CdCl2 with the key residues around iron binding sites.
Co-reporter:Dandan Guo, Bin Zhang and Rutao Liu
RSC Advances 2016 vol. 6(Issue 43) pp:36273-36280
Publication Date(Web):12 Apr 2016
DOI:10.1039/C6RA03122F
NanoAg is almost the most widely used nanoparticle material, and the potential toxicity of nanoAg has aroused widespread concerns regarding its effect on human health. However, because previous studies differed in the preparation of the nanoAg as well as its size, exposure mode and targets, coating and so on, it is difficult to compare the experimental results of these reports and to arrive at an accurate and comprehensive conclusion. Further systematic study of the mechanism underlying the toxicity of nanoAg at the molecular level is necessary and important. Lysozyme was selected to investigate the mechanism underlying the toxicity of nanoAg at the molecular level, and this analysis included transmission electron microscopy, enzyme activity assays, and various spectroscopic methods including fluorescence spectroscopy, synchronous fluorescence spectroscopy, light scattering spectroscopy, UV absorption spectroscopy and circular dichroism spectroscopy. Multi-spectroscopic experiments indicated that nanoAg quenched the fluorescence of lysozyme at the concentrations of nanoAg ranging from 1.0 × 10−6 g mL−1 to 200.0 × 10−6 g mL−1 in the quenching mode of exponential decay using both dynamic and static processes, and was accompanied by complex conformational changes of lysozyme. The interaction between lysozyme and nanoAg inhibited the function of lysozyme. The toxicity of nanoAg was attributed to the smaller lysozyme being surrounded by nanoAg and that nanoAg released silver ions. The results will help provide a strong biophysical basis for research into the toxicity of nanoAg.
Co-reporter:Rui Zhang, Rutao Liu, and Wansong Zong
Journal of Agricultural and Food Chemistry 2016 Volume 64(Issue 34) pp:6630-6640
Publication Date(Web):August 10, 2016
DOI:10.1021/acs.jafc.6b02656
Bisphenol S (BPS) is present in multitudinous consumer products and detected in both food and water. It also has been a main substitute for bisphenol A (BPA) in the food-packaging industry. Yet, the toxicity of BPS is not fully understood. The present study of the toxicity of BPS was divided into two parts. First, oxidative stress, cell viability, apoptosis level, and catalase (CAT) activity in mouse hepatocytes and renal cells were investigated after BPS exposure. After 12 h of incubation with BPS, all of these parameters of hepatocytes and renal cells changed by >15% as the concentration of BPS ranged from 0.1 to 1 mM. Second, the direct interaction between BPS and CAT on the molecule level was investigated by multiple spectral methods and molecular docking investigations. BPS changed the structure and the activity of CAT through binding to the Gly 117 residue on the substrate channel of the enzyme. The main binding forces were hydrogen bond and hydrophobic force.Keywords: bisphenol S; catalase; hepatocyte; oxidative stress; renal cell; spectroscopy;
Co-reporter:Jing Wang, Xinping Yang, Jinhu Wang, Chi Xu, Wandi Zhang, Rutao Liu and Wansong Zong
New Journal of Chemistry 2016 vol. 40(Issue 4) pp:3738-3746
Publication Date(Web):02 Mar 2016
DOI:10.1039/C5NJ02911B
The direct binding of cadmium with lysozyme might cause the structural and functional changes of lysozyme. To better understand the potential toxicity and toxic mechanisms of cadmium, it is of vital importance to characterize the interaction of cadmium with lysozyme. This article investigated the interaction of cadmium chloride (CdCl2) with lysozyme using biophysical methods including the spectroscopic technique, isothermal titration calorimetry (ITC), molecular docking and dynamics simulation studies, and enzyme activity measurements. ITC measurements indicated that the interaction is mainly driven by hydrophobic forces with approximately 3 thermodynamic identical binding sites at 310.15 K. Multi-spectroscopic measurements showed that CdCl2 statically quenched the intrinsic fluorescence of lysozyme, formed complexes with lysozyme and altered its secondary structure. Also, the topology of lysozyme in the presence of CdCl2 was altered. Although complexes were formed, we observed no change of lysozyme activity at low CdCl2 concentration because CdCl2 does not preferentially bind to the active site of lysozyme.
Co-reporter:Mingyang Jing, Wei Song, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2016 Volume 164() pp:103-109
Publication Date(Web):5 July 2016
DOI:10.1016/j.saa.2016.04.008
•This work studied the interacting mechanism of copper with lysozyme.•Copper could spontaneously interact with lysozyme through hydrophobic forces.•Binding of copper to lysozyme was a static quenching process.•Copper induced the conformational changes of lysozyme.•Copper could affect the function of lysozyme by inhibiting its catalytic activity.Although copper is essential to all living organisms, its potential toxicity to human health have aroused wide concerns. Previous studies have reported copper could alter physical properties of lysozyme. The direct binding of copper with lysozyme might induce the conformational and functional changes of lysozyme and then influence the body's resistance to bacterial attack. To better understand the potential toxicity and toxic mechanisms of copper, the interaction of copper with lysozyme was investigated by biophysical methods including multi-spectroscopic measurements, isothermal titration calorimetry (ITC), molecular docking study and enzyme activity assay. Multi-spectroscopic measurements proved that copper quenched the intrinsic fluorescence of lysozyme in a static process accompanied by complex formation and conformational changes. The ITC results indicated that the binding interaction was a spontaneous process with approximately three thermodynamical binding sites at 298 K and the hydrophobic force is the predominant driven force. The enzyme activity was obviously inhibited by the addition of copper with catalytic residues Glu 35 and Asp 52 locating at the binding sites. This study helps to elucidate the molecular mechanism of the interaction between copper and lysozyme and provides reference for toxicological studies of copper.Copper could spontaneously bind lysozyme through hydrophobic forces with a positive ΔH, a favorable ΔS, and a negative ΔG. Binding of copper to lysozyme caused static quenching of the fluorescence, the change of the microenvironment of tryptophan residues and conformational changes of lysozyme. Besides, the lysozyme activity was also inhibited by the addition of copper with catalytic residues Glu 35 and Asp 52 locating at the binding sites. Thus, the direct interaction between copper and lysozyme might induce the conformational and functional alterations of lysozyme.
Co-reporter:Rui Zhang, Lining Zhao, Rutao Liu
Journal of Photochemistry and Photobiology B: Biology 2016 Volume 163() pp:40-46
Publication Date(Web):October 2016
DOI:10.1016/j.jphotobiol.2016.08.011
•The diameter of candida rugose lipase was reduced by bisphenol A.•The predominant force of the binding is hydrophobic interaction.•The interaction changes the secondary structure of Candida rugosa lipase.•Bisphenol A could affect the function of Candida rugosa lipase by increasing its catalytic activity.Bisphenol A is widely used in the manufacture of food packaging and beverage containers and can invade our food and cause contamination. Candida rugose lipase has been a versatile enzyme for biocatalysis and biotransformations to produce useful materials for food, pharmaceutical and flavor. The interactions between bisphenol A and Candida rugosa lipase in vitro were studied by UV–vis, steady-state fluorescence, circular dichroism, synchronous fluorescence, light scattering spectra, molecular docking and enzyme activity assay to better understand the toxicity and toxic mechanisms of bisphenol A. The intrinsic fluorescence of the tryptophan amino acid residue and the secondary structure of the globular protein candida rugose lipase were made use of to thoroughly investigate the structural changes caused by bisphenol A. The results of the fluorescence indicated that bisphenol A interacted with candida rugose lipase and made tryptophan be exposed to a hydrophobic environment. Multi-spectroscopic measurements showed that the addition of bisphenol A increased the intrinsic fluorescence of Candida rugosa lipase, loosened its skeleton structure and changed its secondary structure. Also, the increased activity of Candida rugosa lipase revealed that the position or the structure of the catalytic triad of Candida rugosa lipase may be changed. The molecular docking results showed that bisphenol A bound with the residue Serine 209 which could be another reason for the increased activity of Candida rugosa lipase. Moreover, as can be seen from the results of resonance light scattering and dynamic light scattering, the volume of the Candida rugosa lipase was decreased and the lid may be stripped.
Co-reporter:Mingyang Jing;Wenbao Yan;Xuejie Tan;Yadong Chen
Luminescence 2016 Volume 31( Issue 2) pp:557-564
Publication Date(Web):
DOI:10.1002/bio.2995
Abstract
Human serum albumin (HSA) is the most prominent protein in blood plasma with important physiological functions. Although copper is an essential metal for all organisms, the massive utilization of copper has led to concerns regarding its potential health impact. To better understand the potential toxicity and toxic mechanisms of Cu2+, it is of vital importance to characterize the interaction of Cu2+ with HSA. The effect of Cu2+ on the structure and function of HSA in vitro were investigated by biophysical methods including fluorescence techniques, circular dichroism (CD), time-resolved measurements, isothermal titration calorimetry (ITC), molecular simulations and esterase activity assay. Multi-spectroscopic measurements proved that Cu2+ quenched the intrinsic fluorescence of HSA in a dynamic process accompanied by the formation of complex and alteration of secondary structure. But the Cu2+ had minimal effect on the backbone and secondary structure of HSA at relatively low concentrations. The ITC results indicated Cu2+ interacted with HSA spontaneously through hydrophobic forces with approximately 1 thermodynamic identical binding sites at 298 K. The esterase activity of HSA was inhibited obviously at the concentration of 8 × 10-5 M. However, molecular simulation showed that Cu2+ mainly interacted with the amino acid residues Asp (451) by the electrostatic force. Thus, we speculated the interaction between Cu2+ and HSA might induce microenvironment of the active site (Arg 410). This study has provided a novel idea to explore the biological toxicity of Cu2+ at the molecular level. Copyright © 2015 John Wiley & Sons, Ltd.
Co-reporter:Haoyu Sun;Yingxue Liu;Meng Li;Songlin Han;Xudan Yang
Luminescence 2016 Volume 31( Issue 2) pp:335-340
Publication Date(Web):
DOI:10.1002/bio.2964
Abstract
Chrysoidine is widely used in industry as a type of azo dye, and is sometimes used illegally as a food additive despite its potential toxicity. Human serum albumin (HSA) is one of the most important proteins in blood plasma and possesses major physiological functions. In the present study, the conformational and functional effects of chrysoidine on HSA were investigated by isothermal titration calorimetry (ITC), multiple spectroscopic methods, a molecular docking study and an esterase activity assay. Based on the ITC results, the binding stoichiometry of chrysoidine to HSA was estimated to be 1.5:1, and was a spontaneous process via a single hydrogen bond. The binding of chrysoidine to HSA induced dynamic quenching in fluorescence, and changes in secondary structure and in the microenvironment of the Trp-214 residue. In addition, the hydrogen bond (1.80 Å) formed between the chrysoidine molecule and the Gln-211 residue. The esterase activity of HSA decreased following the addition chrysoidine due to the change in protein structure. This study details the direct interaction between chrysoidine and HSA at the molecular level and the mechanism for toxicity as a result of the functional changes induced by HSA structural variation upon binding to chrysoidine in vitro. This study provides useful information towards detailing the transportation mechanism and toxicity of chrysoidine in vivo. Copyright © 2015 John Wiley & Sons, Ltd.
Co-reporter:Jing Wang, Minglu Hao, Chunguang Liu and Rutao Liu
RSC Advances 2015 vol. 5(Issue 40) pp:31798-31806
Publication Date(Web):30 Mar 2015
DOI:10.1039/C5RA03210E
Cadmium can disturb the intracellular redox state and generate oxidative stress, leading to a variety of diseases. A number of studies have reported that oxidative stress and activation of mitogen-activated protein kinase pathways contributes to the induction or inhibition of apoptosis, depending on the types of cell lines and stimulus. However, the role of oxidative stress and the activation of extracellular signal-regulated kinase (ERK) signaling pathways in the apoptosis of cadmium-treated primary hepatocytes and its underlying mechanism still remain unclear. The present study clearly confirms the important role of oxidative stress in the time-delayed effects of cadmium-induced apoptosis. Rather than direct induction by cadmium, cadmium decreased cell viability by causing oxidative stress mediated-apoptosis after 24 h exposure. This was verified by the persistent elevation of excessive reactive oxygen species and DNA oxidative damage, the decrease in glutathione level and the protection of N-acetyl-L-cysteine (NAC). Furthermore, NAC and PD98059 prevented apoptosis, caspase-3 activation and histone H3 phosphorylation. Also, NAC suppressed Cd-induced activation of the ERK pathway. These results demonstrated that oxidative stress-regulated downstream ERK pathway activation plays a pivotal role in Cd-induced apoptosis of primary hepatocytes. The present study also indicates for the first time that Cd-induced histone H3 phosphorylation is closely associated with activation of the ERK pathway and is involved in oxidative stress-mediated DNA damage and cell death. These results will contribute to a better understanding of the cellular mechanisms of cadmium toxicity and its effects on human health.
Co-reporter:Xiaojing Sun, Wansong Zong, Chunguang Liu, Yang Liu, Canzhu Gao, Rutao Liu
Journal of Luminescence 2015 Volume 161() pp:71-75
Publication Date(Web):May 2015
DOI:10.1016/j.jlumin.2014.12.061
•This work established the binding mode of plasticizers with DNA on molecular level.•The mechanism was explored by fluorescence spectroscopic and molecular docking methods.•There are two kinds of binding mode between DMP, DEP, DBP and DNA, electrostatic and groove.•With the branched chain extension, the binding effect of plasticizers and DNA has been weakened.The interactions of typical plasticizers dimethyl phthalate (DMP), diethyl phthalate (DEP) and dibutyl phthalate (DBP) with calf thymus DNA (ctDNA) were investigated by fluorescence spectroscopic techniques and molecular modeling. Experimental results indicated that the characteristic fluorescence intensity of phthalic acid rose with the increase of DNA concentration; while the characteristic fluorescence intensities of plasticizers decreased with the increase of DNA concentration. Experiments on native and denatured DNA determined that plasticizers interacted with DNA both in groove and electrostatic binding mode. The molecular modeling results further illustrated that there is groove binding between them; hydrogen bonding and Van der Waals interactions were the main forces. With the extension of branched-chains, the binding effects between plasticizers and DNA were weakened, which could be related to the increased steric hindrance.
Co-reporter:Jinhu Wang, Wen Wang, Chunli Liu, Yuliang Zhao, Han Cao, Yongjun Liu and Rutao Liu
RSC Advances 2015 vol. 5(Issue 24) pp:18622-18632
Publication Date(Web):09 Feb 2015
DOI:10.1039/C4RA16652C
Phosphorylation mechanisms of glucose catalyzed by complexes of glucose–ATP–Mg2+–StHK, glucose–ATP–Mg2+–K15A mutant and glucose–ATPγS–Mg2+–StHK have been extensively studied using the quantum mechanical/molecular mechanical (QM/MM) method. Structural analyses show that Mg2+ ion plays a key role in the stabilization of the β-phosphate in the whole catalytic reaction and contributes much to the departure of γ-phosphoryl (or γ-thiophosphoryl) group. Besides, the existence of K15 could also facilitate the stabilization of the β-phosphate directly and influence the binding of γ-phosphate (or γ-thiophosphate) with C6-hydroxyl group indirectly. For each complex, two catalytic processes (the phosphate transfer and proton transfer steps) are studied. The phosphate transfer process is calculated to be the rate-determining step in all three complexes, where the energy barrier of the phosphate transfer is 4.0, 11.7 and 10.9 kcal mol−1 for glucose–ATP–Mg2+–StHK, glucose–ATP–Mg2+–K15A mutant and glucose–ATPγS–Mg2+–StHK complexes, respectively. Both the ATP and ATPγS bound StHK are exothermic, where the catalytic reaction is endothermic for K15A mutant. Calculations suggest that the influence of K15A mutation to the reactive activity is larger than that of the exchange of ATP to ATPγS. We suppose that K15 might play the similar role with those conserved arginine residue in human hexokinase I–IV.
Co-reporter:Lining Zhao;Wei Song;Jing Wang;Yunxing Yan;Jiangwei Chen
Journal of Biochemical and Molecular Toxicology 2015 Volume 29( Issue 12) pp:579-586
Publication Date(Web):
DOI:10.1002/jbt.21731
ABSTRACT
To research the mechanism of dimercaptosuccinic acid coated-superparamagnetic iron oxide nanoparticles (SPION) with human serum albumin (HSA), the methods of spectroscopy, molecular modeling calculation, and calorimetry were used in this paper. The inner filter effect of the fluorescence intensity was corrected to obtain the accurate results. Ultraviolet–visible absorption and circular dichroism spectra reflect that SPION changed the secondary structure with a loss of α-helix and loosened the protein skeleton of HSA; the activity of the protein was also affected by the increasing exposure of SPION. Fluorescence lifetime measurement indicates that the quenching mechanism type of this system was static quenching. The isothermal titration calorimetry measurement and molecular docking calculations prove that the predominant force of this system was the combination of Van der Waals’ force and hydrogen bonds.
Co-reporter:Zehua Yu, Hongwei Liu, Xinxin Hu, Wei Song, Rutao Liu
Journal of Luminescence 2015 159() pp: 312-316
Publication Date(Web):
DOI:10.1016/j.jlumin.2014.10.049
Co-reporter:Fang Hao, Mingyang Jing, Xingchen Zhao, Rutao Liu
Journal of Photochemistry and Photobiology B: Biology 2015 Volume 143() pp:100-106
Publication Date(Web):February 2015
DOI:10.1016/j.jphotobiol.2015.01.003
•The binding stoichiometry of copper ion to catalase is 11.4.•The result of catalase activity on molecular level was consistent with that in the cellular test.•The conformation and function of catalase were changed induced by copper ion.•This study established a combined cellular and molecular toxicity evaluation method.In this research, the binding mechanism of Cu2+ to bovine liver catalase (BLC) was studied by fluorescence spectroscopy, ultraviolet–visible (UV–vis) absorption spectroscopy, circular dichroism (CD) spectroscopy, isothermal titration calorimetry (ITC) and molecular docking methods. The cellar experiment was firstly carried out to investigate the inhibition effect of catalase. During the fluorescence quenching study, after correcting the inner filter effect (IFE), the fluorescence of BLC was found to be quenched by Cu2+. The quenching mechanism was determined by fluorescence lifetime measurement, and was confirmed to be the dynamic mode. The secondary structure content of BLC was changed by the addition of Cu2+, as revealed by UV–vis absorption and CD spectra, which further induces the decrease in BLC activity. Molecular simulation study indicates that Cu2+ is located between two β-sheets and two random coils of BLC near to the heme group, and interacts with His 74 and Ser 113 residues near a hydrophilic area. The decrease of α-helix and the binding of His 74 are considered to be the major reason for the inhibition of BLC activity caused by Cu2+. The ITC results indicate that the binding stoichiometry of Cu2+ to catalase is 11.4. Moreover, the binding of Cu2+ to BLC destroyed H-bonds, which was confirmed by the CD result.Graphical abstract
Co-reporter:Wei Song, Zehua Yu, Xinxin Hu, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2015 Volume 137() pp:286-293
Publication Date(Web):25 February 2015
DOI:10.1016/j.saa.2014.08.037
•The fluorescence of trypsin was quenched by hydrogen peroxide in a static quenching.•Van der Waals forces and hydrogen bonding interactions are main forces during their binding process.•No distinct activity change of trypsin occurs with the concentration of hydrogen peroxide less than 0.12 M.•The toxicity of hydrogen peroxide to trypsin was reported at the molecular level.Studies on the effects of environmental pollutants to protein in vitro has become a global attention. Hydrogen peroxide (H2O2) is used as an effective food preservative and bleacher in industrial production. The toxicity of H2O2 to trypsin was investigated by multiple spectroscopic techniques and the molecular docking method at the molecular level. The intrinsic fluorescence of trypsin was proved to be quenched in a static process based on the results of fluorescence lifetime experiment. Hydrogen bonds interaction and van der Waals forces were the main force to generate the trypsin-H2O2 complex on account of the negative ΔH0 and ΔS0. The binding of H2O2 changed the conformational structures and internal microenvironment of trypsin illustrated by UV–vis absorption, fluorescence, synchronous fluorescence, three-dimensional (3D) fluorescence and circular dichroism (CD) results. However, the binding site was far away from the active site of trypsin and the trypsin activity was only slightly affected by H2O2, which was further explained by molecular docking investigations.Van der Waals forces and hydrogen bonds play the major role in the interaction of trypsin and hydrogen peroxide. The two hydrogen bonds are 1.901 Å and 1.682 Å formed between hydrogen peroxide and the GLN 210.A residue.
Co-reporter:Qianqian Wu, Hao Zhang, Tao Sun, Bin Zhang, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2015 Volume 151() pp:124-130
Publication Date(Web):5 December 2015
DOI:10.1016/j.saa.2015.06.080
•This work investigated the toxic mechanisms of Ag+ to lysozyme.•Ag+ interacted with lysozyme and changed the frame structure of lysozyme to become looser.•Ag+ could spontaneously bind with lysozyme through hydrogen bonding and van der Waals forces.•Ag+ bound to lysozyme at the active site and furtherly influence the function of lysozyme.Silver (Ag) is widely used in human activities, which provides possibilities to distribute in organisms and tissues, resulting in harmful effects on human health. In this work, lysozyme was chosen as the target molecule to study the mechanism of toxic interactions between Ag+ and protein using fluorescence emission spectra, synchronous fluorescence spectra, UV–vis absorption spectra, circular dichroism (CD) spectra, isothermal titration calorimetry (ITC), and enzyme activity assay. The results of fluorescence emission and synchronous fluorescence showed that there were interactions between Ag+ and lysozyme by eliminating the inner filter effect (IFE). Data from UV–vis spectra indicated that the frame structure of lysozyme became looser with Ag+ existent, while the micro-environment of aromatic amino acid residues did not show any significant alteration. CD results suggested that the secondary structure of lysozyme presented a decrease in α-helix contents with the increasing amount of Ag+. ITC results showed Ag+ can spontaneously bind with lysozyme through hydrogen bonding and van der Waals forces with one binding site (Ka = 1.93 × 106). The lysozyme activity was inhibited by Ag+ according to the enzyme activity assay, revealing that Ag+ bound to lysozyme at the active site which resulted in inhibition of lysozyme activity. This work showed that Ag+ can cause damages to the structure and function of lysozyme.Ag+ could spontaneously interact with lysozyme through hydrogen bonds and van der Waals forces with one binding site, resulting in the conformational changes of lysozyme and the decrease of enzyme activity. Thus, Ag+ may influence the structure and function of proteins.
Co-reporter:Jin Guan;Guiliang Liu;Kai Cai;Canzhu Gao
Luminescence 2015 Volume 30( Issue 5) pp:693-698
Publication Date(Web):
DOI:10.1002/bio.2807
Abstract
In order to evaluate the toxicity of multi-walled carbon nanotubes (MWCNTs-COOH) at a molecular level, the effect of MWCNTs-COOH on antioxidant enzyme copper–zinc superoxide dismutase (Cu/ZnSOD) was investigated using fluorescence spectroscopy, UV/vis absorption spectroscopy, circular dichroism (CD) spectroscopy and isothermal titration calorimetry (ITC). By deducting the inner filter effect (IFE), the fluorescence emission spectra and synchronous fluorescence spectra indicated that there were interactions between MWCNTs-COOH and Cu/ZnSOD. Moreover, the microenvironment of the amino acid residues in the enzyme was changed slightly. The UV/vis absorption and CD spectroscopic results showed appreciable conformational changes in Cu/ZnSOD. However, the results of a Cu/ZnSOD activity determination did not show any significant difference. In other words, MWCNTs-COOH has no significant effect on enzyme activity. The ITC results showed that the binding of MWCNTs-COOH to Cu/ZnSOD was a weak endothermic process, indicating that the predominant force of the binding was hydrophobic interaction. Moreover, it was essential to consider the IFE in fluorescence assays, which might affect the accuracy and precision of the results. The above results are helpful in evaluating the oxidative stress induced by MWCNTs-COOH in vivo. Copyright © 2014 John Wiley & Sons, Ltd.
Co-reporter:Zhaozhen Cao, Rutao Liu, Ziliang Dong, Xinping Yang, Yadong Chen
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2015 Volume 136(Part B) pp:601-606
Publication Date(Web):5 February 2015
DOI:10.1016/j.saa.2014.09.073
•The effects of sarafloxacin on Cu/ZnSOD structure and activity were investigated.•Sarafloxacin can bind to Cu/ZnSOD mainly through hydrogen bond and hydrophobic forces.•The binding process tends to be saturated as the concentration of sarafloxacin reaches 4 times of Cu/ZnSOD.•The binding induces structure change in Cu/ZnSOD but does not affect its activity.•Sarafloxacin binds to the surface of β-barrel of Cu/ZnSOD far away from the active site and active site channels.The effect of sarafloxacin to Cu/ZnSOD was evaluated via investigating the change in Cu/ZnSOD structure and the structure basis activity upon sarafloxacin binding. Multi-spectroscopic methods, isothermal titration microcalorimetry (ITC) and molecular docking method were adopted in this study. Sarafloxacin binds to Cu/ZnSOD mainly through hydrophobic and hydrogen bond forces and tends to be saturated as the molar ratio of sarafloxacin to Cu/ZnSOD reaches 4. The binding changed the microenvironment around Tyr and the secondary structure of Cu/ZnSOD but did not affect the activity of Cu/ZnSOD. Molecular docking study revealed that sarafloxacin binds into a hydrophobic area with possibility to form hydrogen bonds with Tyr 108, Asp 25, Pro 100 and Ser 103 of Cu/ZnSOD. The binding area locates on the surface of β-barrel close to the second Greek key loop (GK2) and V-loop but far away from the active site and active site channel of Cu/ZnSOD. These promoted the understanding of the experiment phenomenons. The binding of sarafloxacin does not affect the activity of Cu/ZnSOD should attribute to the binding not to change the microenvironment of Cu/ZnSOD active site and active site channel.The effects of sarafloxacin on Cu/ZnSOD were evaluated via investigating the structure and the structure basis activity changes of Cu/ZnSOD upon sarafloxacin binding using multi-spectroscopic methods, isothermal titration microcalorimetry (ITC) and molecular docking method. Sarafloxacin binds to a hydrophobic area located on the surface of β-barrel and tends to form hydrogen bonds with Tyr 108, Pro 100, Asp 25 and Ser 103 residues around it. The binding of sarafloxacin induces structure change in Cu/ZnSOD but does not affect its activity, which can be attributed to the active site and active site channel of Cu/ZnSOD being far away from the binding site and the microenvironment of them not to be affected.
Co-reporter:Haoyu Sun, Qing Xia, Rutao Liu
Journal of Luminescence 2014 Volume 148() pp:143-150
Publication Date(Web):April 2014
DOI:10.1016/j.jlumin.2013.12.012
Co-reporter:Jin Guan;Jingping Dai;Xingchen Zhao;Chunhua Liu;Canzhu Gao
Journal of Biochemical and Molecular Toxicology 2014 Volume 28( Issue 5) pp:211-216
Publication Date(Web):
DOI:10.1002/jbt.21555
ABSTRACT
The interactions between well-dispersed multiwalled carbon nanotubes (MWCNTs) and catalase (CAT) were investigated. The activity of CAT was inhibited with the addition of MWCNTs. After deducting the inner filter effect, the fluorescence spectra revealed that the tryptophan (Trp) residues were exposed and the fluorescence intensities of CAT increased with the increase in the MWCNTs concentration. At the same time, the environment of the Trp residues became more hydrophobic. The results of UV–vis absorption spectroscopy and CD spectra indicated that the secondary structure of CAT had been changed, and the amino acid residues were located in a more hydrophobic environment. Meanwhile, the UV–vis spectra indicated that the conformation of the heme porphyrin rings was changed. The microenvironment of CAT activity sites may be interfered by MWCNTs. This research showed that MWCNTs could not only contribute to the conformational changes of protein but also change the enzyme function.
Co-reporter:Haoyu Sun;Erqian Cui;Zhigang Tan
Journal of Biochemical and Molecular Toxicology 2014 Volume 28( Issue 12) pp:549-557
Publication Date(Web):
DOI:10.1002/jbt.21597
ABSTRACT
The interactions of N-acetyl-L-cysteine-capped CdTe quantum dots (QDs) with bovine serum albumin (BSA) and bovine hemoglobin (BHb) were investigated by isothermal titration calorimetry (ITC), fluorescence, synchronous fluorescence, fluorescence lifetime, ultraviolet–visible absorption, and circular dichroism techniques. Fluorescence data of BSA–QDs and BHb–QDs revealed that the quenching was static in every system. While CdTe QDs changed the microenvironment of tryptophan in BHb, the microenvironment of BSA kept unchanged. Adding CdTe QDs affected the skeleton and secondary structure of the protein (BSA and BHb). The ITC results indicated that the interaction between the protein (BSA and BHb) and QDs-612 was spontaneous and the predominant force was hydrophobic interaction. In addition, the binding constants were determined to be 1.19 × 105 L mol−1 (BSA–QDs) and 2.19 × 105 L mol−1 (BHb–QDs) at 298 K. From these results, we conclude that CdTe QDs have a larger impact on the structure of BHb than BSA.
Co-reporter:Hao Zhang, Kai Wei, Mengyu Zhang, Rutao Liu, Yadong Chen
Journal of Photochemistry and Photobiology B: Biology 2014 136() pp: 46-53
Publication Date(Web):
DOI:10.1016/j.jphotobiol.2014.04.020
Co-reporter:Haoyu Sun, Bingjun Yang, Erqian Cui, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2014 Volume 132() pp:692-699
Publication Date(Web):11 November 2014
DOI:10.1016/j.saa.2014.04.157
•The interaction between CdTe QDs (QDs-612) and catalase (CAT) is spontaneous.•The predominant force of the binding is hydrophobic interaction.•The interaction changes the secondary structure of CAT.•Tryptophan residues of CAT expose to a hydrophilic environment.•QDs-612 could affect the function of CAT by decreasing its catalytic activity.Quantum dots (QDs) are recognized as some of the most promising semiconductor nanocrystals in biomedical applications. However, the potential toxicity of QDs has aroused wide public concern. Catalase (CAT) is a common enzyme in animal and plant tissues. For the potential application of QDs in vivo, it is important to investigate the interaction of QDs with CAT. In this work, the effect of N-Acetyl-l-cysteine-Capped CdTe Quantum Dots with fluorescence emission peak at 612 nm (QDs-612) on CAT was investigated by fluorescence, synchronous fluorescence, fluorescence lifetime, ultraviolet–visible (UV–vis) absorption and circular dichroism (CD) techniques. Binding of QDs-612 to CAT caused static quenching of the fluorescence, the change of the secondary structure of CAT and the alteration of the microenvironment of tryptophan residues. The association constants K were determined to be K288K = 7.98 × 105 L mol−1 and K298K = 7.21 × 105 L mol−1. The interaction between QDs-612 and CAT was spontaneous with 1:1 stoichiometry approximately. The CAT activity was also inhibited for the bound QDs-612. This work provides direct evidence about enzyme toxicity of QDs-612 to CAT in vitro and establishes a new strategy to investigate the interaction between enzyme and QDs at a molecular level, which is helpful for clarifying the bioactivities of QDs in vivo.Graphical abstractN-Acetyl-l-cysteine-Capped CdTe Quantum Dots with fluorescence emission peak at 612 nm (QDs-612) could interact with catalase (CAT) which leads to conformational and functional changes of the enzyme. The potential toxicity of QDs-612 to CAT was investigated by multi-spectroscopic techniques. Binding of QDs-612 to CAT caused static quenching of the fluorescence, the change of the microenvironment of tryptophan residues and the secondary structure of CAT. The interaction between QDs-612 and CAT was spontaneous through hydrophobic force with about 1:1 stoichiometry. Besides, the CAT activity was also inhibited for the bound QDs-612. This work clarifies the fact that QDs-612 can not only contribute to the conformational changes of CAT but also alter the enzyme function.
Co-reporter:Minglu Hao, Rutao Liu, Hao Zhang, Yating Li, Mingyang Jing
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2014 Volume 125() pp:7-11
Publication Date(Web):5 May 2014
DOI:10.1016/j.saa.2013.12.060
•We established a rapid and accurate detection of GSH content in single cells.•The derivatization of channel-surface proved to be necessary.•Optimized voltage proved to be essential to the injection and separation of samples.•The method is low-cost, time-saving, and eco-friendly.A rapid and accurate detection of glutathione (GSH) content in single cells is important to the early diagnosis and prevention of diseases. A microfluidic system allows the manipulation of trace amounts of reagents and single cells in a simple and cheap glass chip coupled with laser induced fluorescence (LIF) detection. 2,3-Naphthalenedicarboxaldehyde (NDA) was used as the derivatization reagent to label GSH in cells. Microchannel surface derivatization and optimization of injection and separation were investigated in detail, and then the GSH in single mice hepatocyte was separated and detected under optimum conditions with a linear range of 5 × 10−4 M ∼ 5 × 10−3 M and a detection limit of 4.47 × 10−5 M. This study provides a simple and effective method for rapid GSH detection in single cells using few reagents.Graphical abstractThe MCE-LIF system is composed of an intelligent electric driving instrument (IEED), a home-made cross microfluidic chip, a LIF detector (excitation wavelength/emission wavelengths: 473 nm/525 nm) and a data acquisition card. Based on the fluorescence quenching of fluorescinisothiocyate (FITC) by GSH, the internal pipeline of the microfluidic chips, the injection and separation condition were modified and optimized. Our study provided a basic and efficient method and technical support for the detection of GSH content in single hepatocytes.
Co-reporter:Jinhu Wang, Rui Zhang, Rutao Liu, Yongjun Liu
Journal of Molecular Graphics and Modelling 2014 Volume 54() pp:100-106
Publication Date(Web):November 2014
DOI:10.1016/j.jmgm.2014.10.003
•Substrates can be docked into the pocket of E386G mutant.•The rescue mechanism is a concerted process with a calculated energy barrier of 22.6 kcal/mol.•The low barrier suggests that anionic formate might be a good nucleophile.•The external nucleophile has little steric hindrance with pocket residues.In the present study, the quantum mechanical/molecular mechanical (QM/MM) method was used to investigate the rescue mechanism of an E386G mutant as well as the glycosylation mechanism of the wild rice β-d-glucosidase. E386G mutant experiences an asynchronous collaborative process to glycosylate the anionic formate with an energy barrier of 22.6 kcal/mol, while the energy barrier is 25.9 kcal/mol for the wild complex. The low energy barrier of the mutated complex suggests that anionic formate might be a good nucleophile to attack the anomeric carbon atom. Both energy barriers can be lowered when the leaving departure releases from the active site, suggesting that the product release, rather than chemistry, contributes to the rate limiting in BGlu1 mutants. Structure analyses also indicate that the external nucleophile has little steric hindrance with pocket residues and adjusts freely to proceed the rescue mechanism of the mutated complex. Our calculations provide a guide for the selectivity of exogenous nucleophiles in the future study of β-glucosidase.
Co-reporter:Hao Zhang, Yang Liu, Rui Zhang, Rutao Liu, and Yadong Chen
The Journal of Physical Chemistry B 2014 Volume 118(Issue 32) pp:9644-9650
Publication Date(Web):August 6, 2014
DOI:10.1021/jp505565s
Lead exposure could induce endocrine disruption and hormonal imbalance of humans, resulting in detrimental effects on the reproductive system even at low doses. However, mechanisms of lead actions remain unknown. This article investigated lead interactions with human chorionic gonadotropin (HCG) as a conceivable mechanism of its reproductive toxicity by spectroscopic technique, isothermal titration calorimetry (ITC), molecular docking study, and enzyme-linked immunosorbent assay (ELISA). Fluorescence measurements showed that lead acetate dynamically quenched intrinsic fluorescence of HCG through collisional mechanism with the association constant (KSV) in the magnitude of 103 L/mol at the detected temperatures (298, 303, and 310 K). ITC and molecular docking results revealed lead acetate could bind into 5 binding sites of HCG through electrostatic effects (ΔH < 0, ΔS > 0) and hydrophobic forces (ΔH > 0, ΔS > 0). The conformational investigation of HCG by UV–vis absorption spectroscopy, circular dichroism spectroscopy, and ELISA indicated lead acetate changed the secondary structure of HCG by loosening and destruction of HCG skeleton and increasing the hydrophobicity around Tyr residues and resulted in the decreased bioactivities of HCG. This work presents direct interactions of lead with sex hormones and obtains a possible mechanism on lead induced reproductive toxicity at the molecular level.
Co-reporter:Hao Zhang, Yang Liu, Rutao Liu, Chunguang Liu, and Yadong Chen
The Journal of Physical Chemistry B 2014 Volume 118(Issue 51) pp:14820-14826
Publication Date(Web):December 12, 2014
DOI:10.1021/jp511056t
Lead toxicity has been proved to be related with inducing oxidative stress of organisms and causing inactivation of antioxidant enzymes, the mechanism of which remains unknown. This study investigated and compared superoxide dismutase (Cu/Zn SOD) activity inhibited in lead-treated zebrafish livers and explored the mechanism of SOD inactivation by lead at the molecular level using multiple spectroscopic techniques, isothermal titration calorimetric (ITC) measurement, molecular docking study and ICP–AES detection. Results showed lead exposure decreased SOD activities in zebrafish livers due to direct interactions between lead and SOD, resulting in conformational and functional changes of the enzyme. To be specific, Studies at the molecular level indicated that lead bound into the active site channel of SOD, hindered the path of the catalytic substrate (O2–•), damaged its skeleton conformation and secondary structure, and interacted with the enzymatically related residue (Arg 141) through electrostatic forces (ΔH < 0, ΔS > 0), and caused the release of Cu2+ and Zn2+ from the catalytic pocket of SOD. This work shows a correlation between results on organismal and molecular levels, and obtains a possible model hypothesizing mechanisms of lead toxicity using in vitro experiments instead of in vivo ones.
Co-reporter:Hongxu Yang;Wei Song;Mingyang Jing
Journal of Biochemical and Molecular Toxicology 2013 Volume 27( Issue 5) pp:272-278
Publication Date(Web):
DOI:10.1002/jbt.21487
ABSTRACT
By utilizing multispectrosopic techniques, the toxic interaction of 2-aminoanthraquinone (2-AAQ) with calf thymus deoxyribonucleic acid (ctDNA) was investigated in vitro under simulated physiological conditions. The experimental results proved that 2-AAQ has a toxic interaction with ctDNA. The binding capacity of DNA with 2-AAQ is diminishing as the pH value of system increasing in the optimization of experimental condition. Moreover we selected pH 7.4, which is nearly physiological condition to enhance the practical significance. According to the Stern–Volmer equation, the quenching was the static quenching process. And the quenching constant can be derived from the fluorescence quenching spectrogram. Ultraviolet absorption spectra and the change in the fluorescence intensity at different ionic strengths further indicated that there was electrostatic binding between 2-AAQ and ctDNA. The circular dichroism experiment showed that the DNA conformation varied from B to A conformation. The basic group enhanced after 2-AAQ embedding. The double helix is more compact, and the DNA conformation changes. © 2013 Wiley Periodicals, Inc. J BiochemMol Toxicol 27:272-278, 2013; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.21487
Co-reporter:Bingjun Yang;Xiaopeng Hao;Yongzhong Wu
Biological Trace Element Research 2013 Volume 155( Issue 1) pp:150-158
Publication Date(Web):2013 October
DOI:10.1007/s12011-013-9771-z
Quantum dots (QDs) are recognized as some of the most promising candidates for future applications in biomedicine. However, concerns about their safety have delayed their widespread application. Human serum albumin (HSA) is the main protein component of the circulatory system. It is important to explore the interaction of QDs with HSA for the potential in vivo application of QDs. Herein, using spectroscopy and isothermal titration calorimetry (ITC), the effect of glutathione-capped CdTe quantum dots of different sizes on the HSA was investigated. After correction for the inner filter effect, the fluorescence emission spectra and synchronous fluorescence spectra showed that the microenvironment of aromatic acid residues in the protein was slightly changed when the glutathione (GSH)–cadmium telluride (CdTe) QDs was added, and GSH–CdTe QDs with larger particle size exhibited a much higher effect on HSA than the small particles. Although a ground-state complex between HSA and GSH–CdTe QDs was formed, the UV–vis absorption and circular dichroism spectroscopic results did not find appreciable conformational changes of HSA. ITC has been used for the first time to characterize the binding of QDs with HSA. The ITC results revealed that the binding was a thermodynamically spontaneous process mainly driven by hydrophobic interactions, and the binding constant tended to increase as the GSH–CdTe QDs size increased. These findings are helpful in understanding the bioactivities of QDs in vivo and can be used to assist in the design of biocompatible and stable QDs.
Co-reporter:Bingjun Yang, Fang Hao, Jiarong Li, Dongliang Chen, Rutao Liu
Journal of Photochemistry and Photobiology B: Biology 2013 Volume 128() pp:35-42
Publication Date(Web):5 November 2013
DOI:10.1016/j.jphotobiol.2013.08.006
•Chrysoidine inhibits catalase in a non-competitive manner.•No profound conformational change of catalase occurs in the presence of chrysoidine.•ITC results indicate that catalase has two sets of binding sites for chrysoidine.•Chrysoidine is located within the bottleneck in the main channel of catalase.•We report a strategy to investigate the toxicity of chrysoidine at the molecular level.Chrysoidine is an industrial azo dye and the presence of chrysoidine in water and food has become an environmental concern due to its negative effects on human beings. In this work, the interactions between chrysoidine and bovine liver catalase (BLC) were explored. Obvious loss in catalytic activity was observed after incubation of BLC with chrysoidine, and the inhibition effect of BLC was found to be of the non-competitive type. No profound conformational change of BLC occurs in the presence of chrysoidine as revealed by UV–vis absorption, circular dichroism and fluorescence spectroscopy studies. Isothermal titration calorimetry results indicate that catalase has two sets of binding sites for chrysoidine. Further, molecular docking simulations show that chrysoidine is located within the bottleneck in the main channel of the substrate to the active site of BLC, which explain the activity inhibition of BLC by chrysoidine.Graphical abstract
Co-reporter:Jun Chai, Qifei Xu, Jinping Dai, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2013 Volume 105() pp:200-206
Publication Date(Web):15 March 2013
DOI:10.1016/j.saa.2012.12.017
Clenbuterol (CLB) is a kind of β2-adrenergic agonists which was illegally used as feed additives nowadays. The toxic interaction of CLB with trypsin, an important digestive enzyme, was studied in vitro using multi-spectroscopic methods and molecular modeling methods. CLB was proved to bind with trypsin in S1 pocket, forming a complex driven by the dominant force of H-bond. The binding constant was calculated to be 1.79887 × 105 L mol−1 at 289 K and 0.32584 × 105 L mol−1 at 310 K, respectively. The skeleton of trypsin became loosened and unfolded with the amino residues microenvironment changed. The secondary and tertiary structure of trypsin also varied. Molecular modeling studies illustrated specific display of the binding information and explained most of the experiment phenomena. The binding site of CLB induced the fluorescence quenching as well as inhibition of enzyme activity of trypsin. The study confirmed that CLB had potential toxicity on both the structure and function of trypsin and the effects enhanced with the increasing concentration of CLB.Graphical abstractHighlights► Interaction mechanism between clenbuterol and trypsin was investigated. ► Multi-spectroscopic methods and molecular modeling methods were applied. ► The structure and function (enzyme activity) of trypsin were both affected by CLB.
Co-reporter:Jing Lin;Canzhu Gao
Journal of Fluorescence 2013 Volume 23( Issue 5) pp:921-927
Publication Date(Web):2013 September
DOI:10.1007/s10895-013-1217-7
The interaction between Trp-Arg dipetide (WR) and calf thymus DNA (ctDNA) in pH 7.4 Tris-HCl buffer was investigated by multi-spectroscopic techniques and molecular modeling. The fluorescence spectroscopy and UV absorption spectroscopy indicated that WR interacted with ctDNA in a minor groove binding mode and the binding constant was 4.1 × 103. The ionic strength effect and single-stranded DNA (ssDNA) quenching effect further verified the minor groove binding mode. Circular dichroism spectroscopy (CD) was employed to measure the conformation change of ctDNA in the presence of WR. The molecular modeling results illustrated that electrostatic interaction and groove binding coexisted between them and the hydrogen bond and Van der Waals were main acting forces. All the above methods can be widely used to investigate the interaction of peptide with nucleic acids, which contributes to design the structure of new and efficient drugs.
Co-reporter:Xinxin Hu, Zehua Yu, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2013 Volume 108() pp:50-54
Publication Date(Web):May 2013
DOI:10.1016/j.saa.2013.01.072
The toxicity of hydroxyl group of isopropanol to trypsin in aqueous solution was investigated by techniques including UV–visible absorption spectroscopy, fluorescence spectroscopy, circular dichroism (CD) spectroscopy, enzyme activity assay and molecular docking technology. The results of UV–visible absorption spectroscopy and CD spectra indicate that isopropanol could change the secondary structure of trypsin by increasing the content of α-helix and decreasing the content of β-sheet. The tertiary structure of trypsin was also changed owing to the loss of environmental asymmetry of amino acid residues. Isopropanol bound into a hydrophobic cavity on the surface of trypsin by a hydrogen bond located between the hydrogen atom on the hydroxyl of isopropanol and the oxygen atoms on SER 214 and hydrophobic interaction, as the molecular docking results showed. In addition, isopropanol could affect the function of trypsin by increasing its catalytic activity.Graphical abstractIsopropanol could interact with trypsin to form a complex which leads to conformational and functional changes of the enzyme. The potential toxicity of isopropanol was investigated by multi-spectroscopic techniques and molecular modeling study. Molecular modeling has displayed the specific binding site and given information about binding forces and drug-residues distances which are in accordance with conclusions from spectroscopic experiments. Besides, enzyme activity assay has given evidence from the functional aspect to clarify the fact that isopropanol can not only contribute to the conformational changes of protein but also alter the enzyme function.Highlights► The interaction model of isopropanol with trypsin was developed. ► Isopropanol could change the secondary and tertiary structure of trypsin. ► Molecular modeling was used to simulate the binding mode of the protein and ligand. ► Enzyme activity assay showed IPA could increase the catalytic activity of trypsin.
Co-reporter:Zhaozhen Cao, Rutao Liu, Bingjun Yang
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2013 Volume 115() pp:457-463
Publication Date(Web):November 2013
DOI:10.1016/j.saa.2013.06.093
•The interaction mechanism of sarafloxacin with catalase was investigated.•Sarafloxacin binds to catalase mainly through hydrophobic interactions.•The binding alters the conformation of catalase and leads to the inhibition of enzyme activity.•Molecular docking study contributes to a better understanding about the experiment results and the interaction details.The interaction between sarafloxacin and catalase (CAT) was studied by fluorescence spectroscopy, UV–visible absorption spectroscopy, circular dichroism (CD) spectroscopy, isothermal titration microcalorimetry (ITC) and molecular docking method. After deducting the inner filter effect, the fluorescence of CAT was quenched regularly by different concentrations of sarafloxacin. The quenching mechanism was studied by lifetime measurement, and it was proved to be mostly due to static quenching. The formation of sarafloxacin–CAT complex alters the micro-environment of amide moieties and tryptophan (Trp) residues, reduces the α-helix content of the enzyme, changes the peripheral substituents on the porphyrin ring of heme and leads to the inhibition of the enzyme activity. Molecular docking study reveals that sarafloxacin is located between two α-helix of CAT near to Trp 182 and Trp 185 residues, which supports the experimental results and helps to have a more clear understanding about the interaction mechanism. The change in the relative position of His 74 to heme induced by the variation of secondary structure is considered to be the major reason for the reduction of CAT activity. Moreover, sarafloxacin binds into a hydrophobic area of CAT mainly through hydrophobic interactions, which is consistent with the ITC analysis.The interaction mechanism of sarafloxacin with catalase (CAT) was investigated by combining spectroscopic methods, isothermal titration microcalorimetry (ITC) and molecular docking. Sarafloxacin binds into α-helix area (between Leu 158.A–Lys 168.A and Asp 179.A–Glu 190.A) of CAT mainly through hydrophobic forces. The binding alters the secondary structure of the enzyme, induces variation of the peripheral substituents on the porphyrin ring of heme and leads to the inhibition of CAT activity. The change in the relative position of His 74 to heme induced by the alteration on the secondary structure of CAT is considered to be the key factor for the reduction of the enzyme activity.
Co-reporter:Yajing Sun, Fanying Ji, Rutao Liu, Jing Lin, Qifei Xu, Canzhu Gao
Journal of Luminescence 2012 Volume 132(Issue 2) pp:507-512
Publication Date(Web):February 2012
DOI:10.1016/j.jlumin.2011.09.042
The toxic interaction of 2-aminobenzothiazole (2-ABT) with herring sperm DNA (hs-DNA) was investigated in vitro under simulated physiological conditions by multi-spectroscopic techniques and molecular modeling study. The fluorescence spectroscopy and UV absorption spectroscopy indicated that 2-ABT interacted with hs-DNA in a minor groove binding mode. The binding constant and the number of binding sites were 7.2×103 L mol−1 and 0.95, respectively. Circular dichroism spectroscopy (CD) was employed to measure the conformation change of hs-DNA in the presence of 2-ABT, which verified the minor groove binding mode. The molecular modeling results illustrated that 2-ABT tended to bind in the region of rich A–T base pairs through the hydrogen bond between A 18 and amino group of 2-ABT. Sequence specificity was confirmed by comparison on the interactions of 2-ABT with four kinds of bases. This combination of multiple spectroscopic techniques and molecular modeling methods can be widely used in the investigation on the toxic interaction of small molecular pollutants and drugs with biomacromolecules, which contributes to clarify the molecular mechanism of toxicity or side effect in vivo.Highlights► 2-ABT interacts with hs-DNA in minor groove binding mode to form complex in the ratio 1:1. ► DNA conformation changes after binding with 2-ABT. ► ABT–DNA complex is simulated using Autodock and binding free energy is also calculated. ► Besides Van der Waals force, hydrogen bond between adenine and 2-ABT plays an important role. ► Sequence specificity of the binding is verified by the effect of DNA bases on 2-ABT.
Co-reporter:Pengjun Zhang;Ping Lan;Yuening Ma;Yan Gao;Hao Chen;Qian Fang;Wansong Zong
Journal of Biochemical and Molecular Toxicology 2012 Volume 26( Issue 2) pp:54-59
Publication Date(Web):
DOI:10.1002/jbt.20398
Abstract
The interaction of potassium dichromate (Cr(VI)) with bovine serum albumin (BSA) was investigated by fluorescence, synchronous fluorescence, resonance light scattering (RLS), ultraviolet–visible absorption, and circular dichroism (CD) spectroscopies under simulated physiological conditions. The experimental results showed that Cr(VI) could quench the intrinsic fluorescence of BSA following a static quenching process, which indicates the formation of a Cr(VI)–BSA complex. The binding constant (KA) and binding site (n) were measured at different temperatures. The spectroscopic results also revealed that the binding of Cr(VI) to BSA can lead to the loosening of the protein conformation and can change the microenvironment and skeleton of BSA. © 2011 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:54–59 2012; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.20398
Co-reporter:Jing Wang;Pengfei Qin
Journal of Biochemical and Molecular Toxicology 2012 Volume 26( Issue 9) pp:360-367
Publication Date(Web):
DOI:10.1002/jbt.21430
Abstract
Acid yellow 23 (AY23) is a pervasive azo dye used in many fields which is potentially harmful to the environment and human health. This paper studied the toxic effects of AY23 on trypsin by spectroscopic and molecular docking methods. The addition of AY23 effectively quenched the intrinsic fluorescence of trypsin via static quenching with association constants of K290,K = 3.67 × 105 L mol−1 and K310,K = 1.83 × 105 L mol−1. The calculated thermodynamic parameters conformed that AY23 binds to trypsin predominantly via electrostatic forces with one binding site. Conformational investigations indicated the skeletal structure of trypsin unfolded and the microenvironment of tryptophan changed with the addition of AY23. Molecular docking study showed that AY23 interacted with the His 57 and Lys 224 residue of trypsin and led to the inhibition of enzyme activity. This study offers a more comprehensive picture of AY23–trypsin interaction and indicates their interaction may perform toxic effects within the organism. © 2012 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:360–367, 2012; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.21430
Co-reporter:Bingjun Yang;Xiaopeng Hao;Yongzhong Wu
Biological Trace Element Research 2012 Volume 146( Issue 3) pp:396-401
Publication Date(Web):2012 June
DOI:10.1007/s12011-011-9262-z
Due to their unique fluorescent properties, quantum dots present a great potential for biolabelling applications; however, the toxic interactions of quantum dots with biopolymers are little known. The toxic interactions of glutathione-capped CdTe quantum dots with trypsin were studied in this paper using synchronous fluorescence spectroscopy, fluorescence emission spectra, and UV–vis absorption spectra. The interaction between CdTe quantum dots and trypsin resulted in structure changes of trypsin and inhibited trypsin's activity. Fluorescence emission spectra revealed that the quenching mechanism of trypsin by CdTe quantum dots was a static quenching process. The binding constant and the number of binding sites at 288 and 298 K were calculated to be 1.98 × 106 L mol−1 and 1.37, and 6.43 × 104 L mol−1 and 1.09, respectively. Hydrogen bonds and van der Waals' forces played major roles in this process.
Co-reporter:Wansong Zong;Shutao Cao;Qifei Xu
Luminescence 2012 Volume 27( Issue 4) pp:292-296
Publication Date(Web):
DOI:10.1002/bio.1349
ABSTRACT
This article concerns a new and precise strategy for the determination of Cu2+ based on a color reaction and outer filter effects (OFEs). Cu2+ can react with sodium diethyldithiocarbamate trihydrate (DDTC) to form a DDTC–Cu2+ complex with a significant absorption at 447 nm. Being positively correlated with Cu2+, the absorption could be treated as the basis for the determination of Cu2+. When cuvettes containing the complex were fixed in the light path of a fluorescence spectrophotometer, the excitation/emitted light were absorbed by the OFEs, similar to absorption mechanisms of inner filter effects. Under suitable conditions, OFEs from the complex could quantitatively reduce the fluorescence intensities of quinine sulfate and acridine yellow by absorbing the excitation or emission light. Compared with traditional absorption spectroscopy (with a detection limit at 0.9 µmol/L), indirect OEF techniques showed increased sensitivities by about 1 order of magnitude. The strategy could be extended to many different systems where components absorb the excitation wavelength and/or emission wavelength of fluorescers. Copyright © 2011 John Wiley & Sons, Ltd.
Co-reporter:Jing Lin, Rutao Liu, Canzhu Gao
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2012 Volume 97() pp:532-535
Publication Date(Web):November 2012
DOI:10.1016/j.saa.2012.06.027
The interaction between l-Arg and calf thymus DNA (ctDNA) in sodium acetate–acetic acid buffer (pH = 4) was investigated with the use of neutral red (NR) dye as a spectral probe coupled with UV–vis absorption, fluorescence, and circular dichroism (CD) spectroscopy technique. The UV absorption spectroscopy indicated that l-Arg interacted with ctDNA via electrostatic force and the fluorescence enhancing of the DNA–NR system verified the electrostatic interaction. In addition, detectable changes in the CD spectrum of ctDNA in the presence of l-Arg indicated conformational changes in the DNA double helix after interaction with the drug. Docking studies were found to corroborate the experimental results. All these results prove that this drug interacts with ctDNA via an electrostatic binding mode.Graphical abstractl-Arg binds with backbone phosphate groups of the double helix by electrostatic interaction, and it resulted in the conformational and micro-environmental changes of DNA.Highlights► We investigated the interactions of l-Arg with DNA in molecular level. ► The experiments show conformation and activity of ctDNA has been affected by l-Arg. ► Potential links in DNA’s changes of structure and function were found by docking.
Co-reporter:Yue Teng, Hao Zhang and Rutao Liu
Molecular BioSystems 2011 vol. 7(Issue 11) pp:3157-3163
Publication Date(Web):20 Sep 2011
DOI:10.1039/C1MB05271C
4-Aminoantipyrine (AAP) is scarcely administered as an analgesic drug because of the potential side effects. The residue of AAP in the environment possesses a potential threat to human health. In this article, the binding mode of AAP with the important antioxidant enzyme catalase (CAT) was investigated using spectroscopic and molecular docking methods. AAP can interact with CAT to form an AAP–CAT complex. The binding constant, number of binding sites and thermodynamic parameters were measured, which indicated that AAP could spontaneously bind with CAT through electrostatic forces with one binding site. Molecular docking results revealed that AAP bound into the CAT central cavity. UV-visible absorption, synchronous fluorescence and circular dichroism (CD) results provide data concerning conformational and some microenvironmental changes of CAT. Furthermore, the binding of AAP can inhibit CAT activity in erythrocytes. The present study provides direct evidence at a molecular level to show that exposure to AAP could induce changes in the enzyme CAT structure and function. The estimated methods in this work can be applied to characterize interactions of enzyme systems and other pollutants and drugs.
Co-reporter:Yue Teng, Fanying Ji, Chao Li, Zehua Yu, Rutao Liu
Journal of Luminescence 2011 Volume 131(Issue 12) pp:2661-2667
Publication Date(Web):December 2011
DOI:10.1016/j.jlumin.2011.07.005
4-Aminoantipyrine (AAP) is scarcely administered as a kind of analgesic drug because of the side effect. The residue of AAP in the environment poses a potential threat to human health. To evaluate the toxicity of AAP at the protein level, the effects of AAP on lysozyme were investigated using spectroscopic and molecular docking methods. Addition of AAP effectively quenched the intrinsic fluorescence of lysozyme. Static quenching of lysozyme by AAP revealed the formation of complex. After the inner filter effect was eliminated, the number of binding sites, the binding constant and the thermodynamic parameters were measured, and indicated that AAP can spontaneously bind with lysozyme through hydrophobic interactions with one binding site. Molecular docking results revealed that AAP bound into the enzyme active site and interacted with the Trp 62 and Trp 63 residues of lysozyme, which illustrated that the lysozyme activity was inhibited by AAP, in accordance with the results of the lysozyme activity experiment. Furthermore, the binding of AAP can result in demonstrable change of the conformation of lysozyme. This work is helpful for clarifying the molecular toxic mechanism of AAP in vivo.Highlights► This work established the binding mode of AAP with lysozyme in molecular level. ► Mechanism was explored by multiple spectroscopic and molecular docking methods. ► AAP can inhibit lysozyme activity and induce conformational changes in lysozyme.
Co-reporter:Dong Yuan, Zhonglan Shen, Rutao Liu, Canzhu Gao
Journal of Luminescence 2011 Volume 131(Issue 12) pp:2478-2482
Publication Date(Web):December 2011
DOI:10.1016/j.jlumin.2011.06.036
The interaction of La3+ to bovine serum albumin (BSA) has been investigated mainly by fluorescence spectra, UV–vis absorption spectra, and circular dichroism (CD) under simulative physiological conditions. Fluorescence data revealed that the quenching mechanism of BSA by La3+ was a static quenching process and the binding constant is 1.75×104 L mol−1 and the number of binding sites is 1 at 289 K. The thermodynamic parameters (ΔH=−20.055 kJ mol−1, ΔG=−23.474 kJ mol−1, and ΔS=11.831 J mol−1 K−1) indicate that electrostatic effect between the protein and the La3+ is the main binding force. In addition, UV–vis, CD, and synchronous fluorescence results showed that the addition of La3+ changed the conformation of BSA.Highlights► Fluorescence data was used to demonstrate that BSA fluorescence can be quenched by La3+. ► La3+ can bind to BSA with stoichiometric ratio of 1:1. ► BSA–La3+ complex is stabilized mainly by electrostatic effect. ► Synchronous fluorescence and CD data revealed that BSA undergoes conformational changes upon binding to La3+.
Co-reporter:Xingchen Zhao, Feng Sheng, Jianli Zheng, and Rutao Liu
Journal of Agricultural and Food Chemistry 2011 Volume 59(Issue 14) pp:7902-7909
Publication Date(Web):June 22, 2011
DOI:10.1021/jf2011408
Anthocyanins from the purple Solanum tuberosum newly cultivated by the Taian Academy of Agricultural Sciences were extracted and analyzed using high-performance liquid chromatography (HPLC) and UV–vis spectroscopy. Four individual anthocyanins were detected as the major components, and the total anthocyanin content was 273.5 ± 14.3 mg of cyanidin-3-glucoside equiv/100 g of dry seeds. Results of color stability showed that the purple S. tuberosum anthocyanins (PSTAs) are more stable under low pH and temperatures. Heat and general food additives have fine compatibility with PSTAs; however, they are very sensitive with oxidant and reduction. The influence of PSTAs on Cr(VI) targeted to bovine serum albumin (BSA) was also studied. The quenching of BSA fluorescence caused by Cr(VI) could be delayed by PSTAs. UV–vis and circular dichroism (CD) data suggested that PSTAs can protect the secondary and tertiary structures of BSA by probably interacting with Cr(VI) in advance.
Co-reporter:Li Wang, Rutao Liu, Yue Teng
Journal of Luminescence 2011 Volume 131(Issue 4) pp:705-709
Publication Date(Web):April 2011
DOI:10.1016/j.jlumin.2010.11.022
The interaction between Ni2+ and calf thymus DNA (ctDNA) was investigated in simulated physiological buffer (pH 7.4) using the Neutral Red (NR) dye as a spectral probe by UV–vis absorption and fluorescence spectroscopy, as well as CD spectra. The experimental results showed that the conformational changes in DNA helix induced by Ni2+ are the reason for the fluorescence quenching of the DNA–NR system. From the experimental results, conclusion can be drawn that Ni2+ can cause structural changes of ctDNA and bind with DNA by electrostatic interaction. At the same time, the paper proved that conformation changes of DNA can also lead to the fluorescence decrease of DNA–probe systems.Research Highlights► The formation of new non-fluorescence complex or competing binding sites between small molecules and probe are usually the reason of fluorescence quenching in DNA–probe systems. This study proved that conformation changes of DNA induced by Ni2+ can also lead to the fluorescence decrease of DNA–probe systems.
Co-reporter:Dong Yuan;Zhonglan Shen;Zhenxing Chi;Jianhua Zhu
Journal of Biochemical and Molecular Toxicology 2011 Volume 25( Issue 4) pp:263-268
Publication Date(Web):
DOI:10.1002/jbt.20385
Abstract
The interaction of Ce3+ to bovine serum albumin (BSA) has been investigated mainly by fluorescence spectra, UV–vis absorption spectra, and circular dichroism (CD) under simulative physiological conditions. Fluorescence data revealed that the quenching mechanism of BSA by Ce3+ was a static quenching process, the binding constant is 6.70 × 105, and the number of binding site is 1. The thermodynamic parameters (ΔH = −29.94 kJ mol−1, ΔG = −32.38 kJ mol−1, and ΔS = 8.05 J mol−1 K−1) indicate that electrostatic effect between the protein and the Ce3+ is the main binding force. In addition, UV–vis, CD, and synchronous fluorescence results showed that the addition of Ce3+ changed the conformation of BSA. © 2011 Wiley Periodicals, Inc. J Biochem Mol Toxicol 25:263–268, 2011; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.20385
Co-reporter:Yihong Liu;Yue Mou;Guangjun Zhou
Journal of Biochemical and Molecular Toxicology 2011 Volume 25( Issue 2) pp:95-100
Publication Date(Web):
DOI:10.1002/jbt.20364
Abstract
The mechanism of formaldehyde–protein interactions was investigated by determining the effects of formaldehyde on the common protein bovine serum albumin (BSA). The effects at the molecular level were determined by fluorescence, ultraviolet absorption, and circular dichroism (CD) spectrometry. Formaldehyde could decrease the amount of alpha-helix, leading to loosening of the protein skeleton. In the loose structure, internal amino acids are exposed and the characteristic fluorescence of BSA is obviously quenched. The spectroscopic results reveal that formaldehyde exposure induces changes in the microenvironment and conformation of serum albumin, which could lead to toxic effects on the organism. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 25:95–100, 2011; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.20364
Co-reporter:Zhenxing Chi and Rutao Liu
Biomacromolecules 2011 Volume 12(Issue 1) pp:
Publication Date(Web):December 13, 2010
DOI:10.1021/bm1011568
Because of the widely usage of the veterinary drug tetracycline (TC), its residue exist extensively in the environment (e.g., animal food, soils, surface water, and groundwater) and can enter human body, being potential harmful. Human serum albumin (HSA) is a major transporter for endogenous and exogenous compounds in vivo. The aim of this study was to examine the interaction of HSA with TC through spectroscopic and molecular modeling methods. The inner filter effect was eliminated to get accurate binding parameters. The site marker competition experiments revealed that TC binds to site II (subdomain IIIA) of HSA mainly through electrostatic interaction, illustrated by the calculated negative ΔH° and ΔS°. Furthermore, molecular docking was applied to define the specific binding sites, the results of which show that TC mainly interacts with the positively charged amino acid residues Arg 410 and Lys 414 predominately through electrostatic force, in accordance with the conclusion of thermodynamic analysis. The binding of TC can cause conformational and some microenvironmental changes of HSA, revealed by UV−visible absorption, synchronous fluorescence, and circular dichroism (CD) results. The accurate and full basic data in the work is beneficial to clarifying the binding mechanism of TC with HSA in vivo and understanding its effect on protein function during the blood transportation process.
Co-reporter:Hao Zhang, Rutao Liu, Zhenxing Chi, Canzhu Gao
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2011 Volume 78(Issue 1) pp:523-527
Publication Date(Web):January 2011
DOI:10.1016/j.saa.2010.11.021
In this work, the toxic influence of metallic ions (Na+, Cu2+, Al3+) on the serum albumin were studied by fluorescence, resonance light scattering (RLS), synchronous fluorescence, UV–vis absorption and circular dichroism (CD) spectroscopy. The experimental results indicated that ion electric charge is not the main factor affecting the structure of bovine serum albumin (BSA). Na+ made the structure of BSA tighter and hydrophobicity enhanced, which improved fluorescence intensity, while Cu2+ could react with some functional groups of BSA, making the structure of BSA looser, so that the internal hydrophobic groups such as tryptophan (Trp) and other aromatic residues were gradually exposed. When we observed them with fluorescence spectra, we found fluorescence quenching with increasing Cu2+ dose. Al3+ is shown as little significant influence on the BSA, but BSA was found to aggregate with the dose of Al3+ by means of RLS because of the hydrolysis and ion strength effect of Al3+. The results also proved normal saline could keep lives healthy and good-working as a biological humour, however, heavy metals made harmful effects to the body when they exceeded the minimal effect level (MEL), such as Cu2+ chosen in our work.
Co-reporter:Wansong Zong, Rutao Liu, Changying Guo, Feng Sun
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2011 Volume 78(Issue 5) pp:1581-1586
Publication Date(Web):May 2011
DOI:10.1016/j.saa.2011.02.004
Biomarkers held both incredible application and significant challenge in probing the oxidation mechanisms of proteins under oxidative stress. Here, mass spectrometry (MS) coupled with liquid chromatography (LC) was applied to establish a new pipeline to probe the oxidation sites and degrees of horse cytochrome c (HCC) with its oxidative products serving as the biomarkers. Samples of native and UV/H2O2 oxidized HCCs were digested by trypsin and subjected to biomarker discovery using LC/MS and tandem mass spectrometry (MS/MS). Experiment results proved that the main oxidation sites were located at Cys14, Cys17, Met65 and Met80 residues in peptides C14AQC(heme)HTVEK22, C14AQCHTVEK22, E60ETLMEYLENPKK73, M80IFAGIK86 and M80IFAGIKK87. Quantitative analysis on the oxidized peptides showed the oxidation degrees of target sites had positive correlations with extended oxidation dose and controlled by residues types and their accessibility to solvent molecules. Being able to provide plentiful information for the oxidation sites and oxidation degrees, the identified oxidized products were feasibility biomarkers for HCC oxidation, compared with the conventional protein carbonyl assay.Graphical abstractThe main oxidation sites of horse cytochrome c susceptible to oxidative stress were located at Cys14, Cys17, Met65 and Met80 residues and oxidation to these sites was controlled by their accessibility to solvent molecules. These oxidative products were valid biomarkers for protein oxidation.Research highlights► New biomarker pipeline for cytochrome c oxidation is established based on LC and MS. ► Residues sensitive to oxidative stress are located at Cys14, Cys17, Met65 and Met80. ► Oxidation to these sites is controlled by residue type and residue location. ► Dose-effect experiments verify they are valid biomarkers for cytochrome c oxidation.
Co-reporter:Chai Jun, Yan Xue, Rutao Liu, Meijie Wang
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2011 Volume 79(Issue 5) pp:1406-1410
Publication Date(Web):September 2011
DOI:10.1016/j.saa.2011.04.076
The toxic interaction of methanol, ethanol and propanol with bovine hemoglobin (BHb) at protein molecular level was studied by resonance light scattering (RLS), fluorescence, ultraviolet–visible absorption (UV–vis) and circular dichroism (CD) techniques. The experimental results showed that the three alcohols all had toxic effects on BHb and the effects increased along with the increasing alcohol dose. The results of RLS and fluorescence spectroscopy showed that alcohols can denature BHb. They changed the microenvironment of amino acid residues and led to molecular aggregation. The decreasing order of the influence is propanol, ethanol and methanol. The results of UV–vis and CD spectra revealed that alcohols led to conformational changes of BHb, including the loosening of the skeleton structure and the decreasing of α-helix in the second structure. The changes generated by propanol were much larger than those by methanol and ethanol.Graphical abstractWith the adding of alcohol, the Trp residues buried were exposed, leading to the fluorescence quenching. Meanwhile, the BHb molecules were denatured and aggregated to wrap the Trp residues up, resulting in the fluorescence intensity increasing. The two opposite effects competed and at last, the intensity increased.Highlights► The interaction of methanol, ethanol and propanol with bovine hemoglobin was investigated. ► Toxic effects of the three alcohols increased along with the alcohol dose. ► The degeneration level of BHb is related to the length of carbon chain of alcohol. ► The decreasing order of the influence is propanol, ethanol and methanol.
Co-reporter:Yue Mu, Jing Lin, Rutao Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2011 Volume 83(Issue 1) pp:130-135
Publication Date(Web):December 2011
DOI:10.1016/j.saa.2011.07.092
The toxicity of sodium benzoate to trypsin was investigated by fluorescence spectroscopy, synchronous fluorescence spectroscopy, UV–visible absorption spectroscopy and circular dichroism (CD) spectroscopy under mimic physiological conditions. Sodium benzoate could unfold trypsin by decreasing the β-sheet structure, which leads to more exposure of internal amino acid groups and the obvious intrinsic fluorescence quenching with the rising concentration of sodium benzoate. The results of spectroscopic measurements indicated that sodium benzoate changed the internal microenvironment of trypsin and induced the alteration of the whole molecule, which were performed toxic effects on the organism. Trypsin and sodium benzoate interacted with each other to produce a substance by van der Waals forces and hydrogen bond, the model of which was shown by AutoDock software.Graphical abstractHighlights► Trypsin can interact with sodium benzoate and generate a new-formed complex. ► The main force for the complex are hydrogen bond and van der Waals’ forces. ► We model the structure of the new complex. ► Sodium benzoate changes the structure of trypsin.
Co-reporter:Xingren Pan, Rutao Liu, Pengfei Qin, Li Wang, Xingchen Zhao
Journal of Luminescence 2010 Volume 130(Issue 4) pp:611-617
Publication Date(Web):April 2010
DOI:10.1016/j.jlumin.2009.11.004
Azo dyes, which are common in the environment, can be toxic to various organisms. In order to determine the molecular mechanism of acid yellow 11(AY) toxicity, we studied the effect of AY exposure to the common protein bovine serum albumin (BSA) by several spectroscopic techniques including fluorescence spectroscopy, ultraviolet spectrophotometry (UV) and circular dichroism (CD). It could be concluded from the fluorescence spectra that the quenching effect of BSA by AY was mainly due to complex formation which was unrelated to the absorption of AY. The enthalpy change (ΔH) and entropy change (ΔS) were found to be −21.94 kJ/mol and 30.04 Jmol-1 K-1, respectively. The results confirm that electrostatic attraction was the predominant intermolecular force between BSA and AY. Furthermore, the binding distance (r) between AY and the inner tryptophan residue of BSA was determined to be 3.541 nm on the basis of Forster theory of non-radiative energy transfer. In addition, the conformational changes of BSA in the presence of AY were also analyzed by UV and CD. These results indicated that AY could interact with BSA by complex formation, which also affected the structure of BSA.
Co-reporter:Pengfei Qin, Rutao Liu, Xingren Pan, Xiaoyan Fang and Yue Mou
Journal of Agricultural and Food Chemistry 2010 Volume 58(Issue 9) pp:5561-5567
Publication Date(Web):April 19, 2010
DOI:10.1021/jf100412q
Perfluoroalkyl acids (PFAAs), an emerging class of globally environmental contaminants, pose a great threat to humans with wide exposure from food and other potential sources. To evaluate the toxicity of PFAAs at the protein level, the effects of three PFAAs on bovine serum albumin (BSA) were characterized by fluorescence spectroscopy, synchronous fluorescence spectroscopy, and circular dichroism (CD). On the basis of the fluorescence spectra and CD data, we concluded that perfluoropentanoic acid (PFPA) had little effect on BSA. However, perfluorooctanoic acid (PFOA) and perfluorodecanoic acid (PFDA) exhibited remarkable fluorescence quenching, which was attributed to the formation of a moderately strong complex. The enthalpy change (ΔH) and entropy change (ΔS) indicated that van der Waals forces and hydrogen bonds were the dominant intermolecular forces in the binding of PFAAs to BSA. Furthermore, the BSA conformation was slightly altered in the presence of PFOA and PFDA, with a reduction of α helix. These results indicated that PFAAs indeed impact the conformation of BSA, and PFAAs with longer carbon chains were more toxic, especially at lower concentrations.
Co-reporter:Zhenxing Chi, Rutao Liu, Yue Teng, Xiaoyan Fang, and Canzhu Gao
Journal of Agricultural and Food Chemistry 2010 Volume 58(Issue 18) pp:10262-10269
Publication Date(Web):August 27, 2010
DOI:10.1021/jf101417w
The residue of the widely used veterinary drug oxytetracycline (OTC) in the environment (e.g., animal food, soils, surface water, and groundwater) is potentially harmful. Knowledge of its binding to proteins contributes to the understanding of its toxicity in vivo. This work establishes the binding mode of OTC with bovine serum albumin (BSA) under physiological conditions by spectroscopic methods and molecular modeling techniques. The inner filter effect was eliminated to get accurate data (binding parameters). On the basis of the thermodynamic results and site marker competition experiments, it was considered that OTC binds to site II (subdomain IIIA) of BSA mainly by electrostatic interaction. Furthermore, using the BSA model established with CPHmodels, molecular docking and some other molecular modeling methods were applied to further define that OTC interacts with the Arg 433, Arg 436, Ala 429, and Pro 516 residues of BSA. In addition, UV−visible absorption, synchronous fluorescence, and circular dichroism (CD) results showed that the binding of OTC can cause conformational and some microenvironmental changes of BSA. The work provides accurate and full basic data for clarifying the binding mechanisms of OTC with BSA in vivo and is helpful for understanding its effect on protein function during its transportation and distribution in blood.
Co-reporter:Rutao Liu;Pengfei Qin;Li Wang;Xingchen Zhao;Yihong Liu;Xiaopeng Hao
Journal of Biochemical and Molecular Toxicology 2010 Volume 24( Issue 1) pp:66-71
Publication Date(Web):
DOI:10.1002/jbt.20314
Abstract
The toxic effects of ethanol on bovine serum albumin (BSA) were measured by resonance light scattering (RLS), fluorescence spectroscopy, ultraviolet spectrophotometry (UV), circular dichroism (CD), and transmission electron microscopy (TEM). The results indicated that ethanol had toxic effects on BSA, which led to protein denaturation and the effects increased with the ethanol dose. By means of RLS, BSA was found to aggregate in the presence of ethanol and particles smaller than 100 nm were observed from TEM. The fluorescence spectra showed that the intensity of the characteristic peak of BSA decreased and blue shifted, because of changes in the BSA skeleton structure, as well as alteration of the microenvironment of tryptophan (Trp) residues. The conformation changes of BSA were also shown by UV and CD spectrometry. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:66–71, 2010; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20314
Co-reporter:Yajing Sun;Zhenxing Chi;Pengfei Qin;Xiaoyan Fang ;Yue Mou
Journal of Biochemical and Molecular Toxicology 2010 Volume 24( Issue 5) pp:323-329
Publication Date(Web):
DOI:10.1002/jbt.20341
Abstract
The toxic interaction of melamine with herring sperm DNA (hs-DNA) was investigated by using fluorescence and UV–vis absorption spectra techniques. The experimental results showed that the toxic interaction between melamine and hs-DNA occurred. Fluorescence quenching experiments indicated the existence of electrostatic binding between melamine and hs-DNA. The binding constants KA and the binding site numbers were calculated by means of the Stern–Volmer equation and were 9.8 × 104 L mol−1 and 1.3, respectively. Both the results of fluorescence spectra and UV–vis absorption spectra verified that there are electrostatic binding between melamine and hs-DNA. The possibility in the presence of a classical intercalation binding mode could be ruled out by using DNA unwinding experiments. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:323–329, 2010; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.20341
Co-reporter:Feng Sun, Wansong Zong, Rutao Liu, Jun Chai, Ying Liu
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2010 Volume 76(Issue 2) pp:142-145
Publication Date(Web):July 2010
DOI:10.1016/j.saa.2010.03.002
The fluorescence characteristics of protein molecules are mainly due to their tryptophan (Trp), tyrosine (Tyr) and phenylalanine (Phe) residues, among which tryptophan is the most important. Studying the influence of the micro-environment on tryptophan fluorescence can give us direct and convincing evidence for changes of protein structure and function. In this paper, fluorescence spectroscopy was used to evaluate the changes of tryptophan fluorescence under a variety of micro-environmental conditions (temperature, pH, polarity, presence of surfactants and oxidants) and the mechanisms responsible. This study not only presents more direct evidence to explain how and why the protein fluorescence spectra change, but also provides a new method for analyzing the effect of environmental changes on protein function.
Co-reporter:Li Wang, Rutao Liu, Zhenxing Chi, Bingjun Yang, Pengjun Zhang, Meijie Wang
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2010 Volume 76(Issue 2) pp:155-160
Publication Date(Web):July 2010
DOI:10.1016/j.saa.2010.03.006
The toxic interaction between Ni2+ and bovine hemoglobin (BHb) was investigated using fluorescence spectroscopy, synchronous fluorescence spectroscopy, ultraviolet–visible (UV–vis) absorption spectroscopy and circular dichroism spectroscopy (CD) under simulated physiological conditions. The experimental results showed that both dynamic and static quenching occurred simultaneously when Ni2+ quenched the fluorescence of BHb. The binding site number n, apparent binding constant Ka and corresponding thermodynamic parameters were measured at different temperatures. There was formation of Ni–BHb complex, but the binding between Ni2+ and BHb was not strong. The process of the formation of Ni–BHb complex was a spontaneous interaction procedure in which electrostatic interaction played a major role. In addition, UV–vis and CD results showed that the addition of Ni2+ changed the conformation of BHb.
Co-reporter:Zhenxing Chi, Rutao Liu, Xingren Pan, Yue Teng, Hao Qin, Jianhua Zhu, Xiaopeng Hao
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2010 Volume 75(Issue 1) pp:177-182
Publication Date(Web):January 2010
DOI:10.1016/j.saa.2009.10.009
The toxic interaction of the azo dye-chrysoidine hydrochloride combined with cetyltrimethyl ammonium bromide (CTMAB) in living tissue was studied in vitro. The absorption spectrum, resonance light scattering (RLS), circular dichroism (CD) and transmission electron microscopy (TEM) results showed that the toxicity of chrysoidine hydrochloride itself to calf thymus DNA (ct-DNA) is weak, while the chrysoidine hydrochloride–CTMAB combined pollution showed obvious toxic interaction with ct-DNA. The chrysoidine hydrochloride–CTMAB combined contamination can interact with ct-DNA to form an ion-associated complex through electrostatic and hydrophobic forces. The conformation of DNA was changed in the interaction process to show toxic. The experimental results showed that the combination of chrysoidine hydrochloride–CTMAB has higher toxicity to ct-DNA than either chrysoidine hydrochloride or CTMAB individually, and the combined pollution showed a strong toxic co-effect at a dose of 3.0 × 10−4 mol L−1 chrysoidine hydrochloride and 1.6 × 10−5 mol L−1 CTMAB.
Co-reporter:Zhenxing Chi, Rutao Liu, and Hao Zhang
Biomacromolecules 2010 Volume 11(Issue 9) pp:
Publication Date(Web):August 3, 2010
DOI:10.1021/bm100633h
Oxytetracycline (OTC) is a kind of widely used veterinary drugs. The residue of OTC in the environment (e.g., animal food, soils, surface, and groundwater) is potentially harmful. In this article, the binding mode of OTC with trypsin was investigated using spectroscopic and molecular docking methods. OTC can interact with trypsin with one binding site to form OTC−trypsin complex, resulting in inhibition of trypsin activity and change of the secondary structure and the microenvironment of the tryptophan residues of trypsin. After elimination of the inner filter effect, the association constant, K, was calculated to be K290K = 1.36 × 105 L mol−1, K298K = 7.30 × 104 L mol−1, and K307K = 3.58 × 104 L mol−1 at three different temperatures. The calculated thermodynamic parameters (negative values of ΔH○ and ΔS○) indicated that van der Waals interactions and hydrogen bonds play a major role during the interaction. The molecular docking study revealed that OTC bound into the S1 binding pocket, which illustrates that the trypsin activity was competitively inhibited by OTC, in accordance with the conclusion of the trypsin activity experiment. This work establishes a new strategy to probe the toxicity of OTC and contributes to clarify its molecular mechanism of toxicity in vivo. The combination of spectroscopic and molecular docking methods in this work can be applied to investigate the potential enzyme toxicity of other small organic pollutants and drugs.
Co-reporter:Feng Sun, Rutao Liu, Wansong Zong, Yanmin Tian, Meijie Wang and Pengjun Zhang
The Journal of Physical Chemistry B 2010 Volume 114(Issue 19) pp:6350-6353
Publication Date(Web):April 23, 2010
DOI:10.1021/jp911772q
The cleavage processes of protonated peptides in mass spectrometry, described in the mobile proton model, are charge-directed and depend on the charge distribution around the cleavage sites. Previous studies experimentally verified the mobile proton model by changing peptide sequences. In this study, oxidation was applied to change the charge distribution of peptides, but the sequence was retained. Tandem mass spectrometry (MS/MS) and quantum chemical calculations at the B3LYP/6-31G(d) level were used to test the validity of the mobile proton model. The results showed prominent differences of peptide fragmentation efficiency caused by the charge distribution produced by various oxidation levels. Fragmentation efficiency curves coupled with the relative intensities of the fragments indicated that the cleavage of the peptide Ala-Arg-Arg-Ala (ARRA) became more and more difficult as O atoms were added. The relative charge ratios between C and N atoms in the amide bonds decreased with the increase of oxidation extent, suggesting that oxidation resulted in protons moving away from the amide bonds. The combined methods proposed here provide a unique approach to substantiate and refine the mobile proton model for peptide fragmentation.
Co-reporter:Feng Sun;Wansong Zong
Journal of The American Society for Mass Spectrometry 2010 Volume 21( Issue 11) pp:1857-1862
Publication Date(Web):2010 November
DOI:10.1016/j.jasms.2010.06.022
The influence of charge state on the peptide dissociation behavior in tandem mass spectrometry (MS/MS) is worthy of discussion. Comparative studies of singly- and doubly-protonated peptide molecules are performed to explore the effect and mechanism of charge state on peptide fragmentation. In view of the charge-directed cleavage of protonated peptides described in the mobile proton model, radiolytic oxidation was applied to change the charge distribution of peptides but retain the sequence. Experimental studies of collision energy-dependent fragmentation efficiencies coupled with quantum chemical calculations indicated that the cleavage of ARRA and its side-chain oxidation products with oxygen atoms added followed a trend that doubly-protonated peptides fragment more easily than singly-protonated forms, while the oxidation product with the guanidine group deleted showed the opposite trend. By analyzing the charge distribution around the amide bonds, we found that the relative charge ratios between C and N atoms (QC/QN) in the amide bonds provided a reasonable explanation for peptide fragmentation efficiencies. An increase of the QC/QN value of the amide bond means that a peptide fragments more easily, and vice versa. The results described in this paper provide an experimental and calculation strategy for predicting peptide fragmentation efficiency.
Co-reporter:Yanmin Tian, Rutao Liu, Wansong Zong, Feng Sun, Meijie Wang, Pengjun Zhang
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2010 Volume 75(Issue 2) pp:908-911
Publication Date(Web):February 2010
DOI:10.1016/j.saa.2009.12.034
Hydroxyl radicals generated from Fenton reaction were used to damage the angiotensin. The oxidative damage degree and sites of peptides were measured by HPLC–MS and MS/MS. Experimental results proved that the oxidative damage degree increased with longer reaction time. The results also showed that the side chains of phenylalanine and tyrosine in angiotension can be attacked by hydroxyl radicals to form the oxidative products. A new strategy was established to monitor the oxidative degree and sites of peptides and laid the foundation for protein oxidation. This method can be used to investigate the mechanism of protein oxidative damage caused by oxidative stress which is induced by environmental pollutants and physiological activities. There will also be a wide application in the research of pathogenesis of some disease related to oxidative stress.
Co-reporter:Xingchen Zhao, Rutao Liu, Zhenxing Chi, Yue Teng and Pengfei Qin
The Journal of Physical Chemistry B 2010 Volume 114(Issue 16) pp:5625-5631
Publication Date(Web):April 7, 2010
DOI:10.1021/jp100903x
Bovine serum albumin (BSA) nonspecifically binds to well-dispersed multiwalled carbon nanotubes (MWCNTs), forming a stable bioconjugate. After accounting for the inner filter effect, we found the fluorescence intensity of BSA was quenched by MWCNTs in static mode, which was authenticated by lifetime measurements and Stern−Volmer calculations. The thermodynamic parameters ΔG°, ΔS°, and ΔH° were −9.67 × 103 + 2.48 × 103 ln λ J·mol−1, 41.0 − 0.828 ln λ J·mol−1·K−1, and 7.30 × 103 + 2.23 × 103 ln λ J·mol−1 (λ < 1 × 10−4), respectively, which shows a spontaneous and electrostatic interaction. Scatchard analysis and UV−visible results provide statistical data concerning changes in the microenvironment of amide moieties in response to different doses of MWCNTs, revealing different behavior of the BSA molecules. The absorption spectra also show that the tertiary structure of the protein was partially destroyed. The content of secondary structure elements of BSA was changed by the tubes. This work elucidates the interaction mechanism of BSA and MWCNTs from a spectroscopic angle.
Co-reporter:Zhenxing Chi, Rutao Liu, Lingzi Zhao, Pengfei Qin, Xingren Pan, Feng Sun, Xiaopeng Hao
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2009 Volume 72(Issue 3) pp:577-581
Publication Date(Web):April 2009
DOI:10.1016/j.saa.2008.10.044
The environmental genotoxic behavior of silver nanoparticles (nanoAg) combined with the detergent cetylpyridine bromide (CPB) was examined in vitro. The experimental results showed that the genotoxicity of nanoAg itself is weak, but nanoAg shows obvious genotoxicity after combined with CPB. The combined materials have a strong coeffect on calf thymus DNA (ctDNA) at a concentration of 3.3 × 10−6 g mL−1 nanoAg and 6.0 × 10−6 mol L−1 CPB. After the addition of ctDNA to the nanoAg–CPB system, the particles are scattered and the diameter decreases, which indirectly reveal that nanoAg–CPB has genotoxicity.
Co-reporter:Wansong Zong;Lingzi Zhao;Yanmin Tian;Dong Yuan;Canzhu Gao
Amino Acids 2009 Volume 37( Issue 4) pp:559-564
Publication Date(Web):2009 October
DOI:10.1007/s00726-008-0173-z
In this paper, oxidative damage to the cysteine (CySH) side-chain on a glassy carbon electrode (GCE) was investigated. Voltammetric studies show that there are three anodic peaks for the oxidation of CySH, which arise from (1) the oxidation of the –SH side-chain, forming cystine (0.71 V, vs. SCE) and (2) CySOxH, x = 2, 3 (0.98 V vs. SCE), and (3) the oxidation of the amino acid carboxyl group (around 1.51 V vs. SCE). The influence of dissolved oxygen, pH, scan rate, scan time, temperature and CySH concentration were investigated and the oxidative mechanism proposed. The peaks near 0.71 and 0.98 V are the promising candidates for measuring the oxidation of CySH on the GCE. This paper provides a new strategy for researching oxidative damage of amino acids, sulfur-containing peptides and proteins.
Co-reporter:Rutao Liu, Wansong Zong, Kaikai Jin, Xuetang Lu, Jianhua Zhu, Lijun Zhang, Canzhu Gao
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2008 Volume 70(Issue 1) pp:198-200
Publication Date(Web):June 2008
DOI:10.1016/j.saa.2007.07.037
By using the techniques of resonance light scattering (RLS) and absorption spectra, we studied the toxicosis and detoxifcation mechanism of anionic surfactant SDBS and cationic surfactant CTMAB targeted to bovine serum albumin (BSA). Small quantity of CTMAB combines with SDBS–BSA complex to form SDBS–BSA–CTMAB complex and the IRLS of system enhanced greatly. With the cumulation of quantity, CTMAB captures SDBS from SDBS–BSA complex by electrostatic attraction and CTMAB–SDBS complex forms, meanwhile BSA automatically frees, which is corresponding to the toxicosis and detoxifcation process. Absorption experiment validates that SDBS induces the denaturalization of BSA and CTMAB facilitates the refolding of this protein.
Co-reporter:Rutao Liu, Wansong Zong, Canzhu Gao, Zhenxing Chi, Lijun Zhang
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2007 Volume 68(Issue 1) pp:150-155
Publication Date(Web):September 2007
DOI:10.1016/j.saa.2006.10.057
This article concerns a new and clean strategy for the determination of Cu2+ in electroless copper plating baths with differential spectrophotometry. With this strategy, the problem of too high absorbance of Cu2+ under plating conditions has been solved. We investigated the influence of copper sulfate, sodium hypophosphite, nickel sulfate, sodium citrate, polyglycol, temperature and pH on the absorption spectrum of Cu2+ in electroless copper plating baths. Five grams per litre of CuSO4·5H2O solution was selected as the reference solution. Experimental results prove that, this strategy has the merits of fast and high accuracy compared to the traditional techniques. Linearly dependent coefficient of the working curve is 0.9999 and the components in the formula have no obvious effect on the detection of Cu2+ under experimental conditions. Therefore, we can directly move solutions from the EC plating baths for detection, after that the sample can still go back to the baths without any pollution from the plating process to the environment.
Co-reporter:Feng Sun, Wansong Zong, Rutao Liu, Meijie Wang, Pengjun Zhang, Qifei Xu
Journal of the American Society for Mass Spectrometry (November 2010) Volume 21(Issue 11) pp:1857-1862
Publication Date(Web):1 November 2010
DOI:10.1016/j.jasms.2010.06.022
The influence of charge state on the peptide dissociation behavior in tandem mass spectrometry (MS/MS) is worthy of discussion. Comparative studies of singly- and doubly-protonated peptide molecules are performed to explore the effect and mechanism of charge state on peptide fragmentation. In view of the charge-directed cleavage of protonated peptides described in the mobile proton model, radiolytic oxidation was applied to change the charge distribution of peptides but retain the sequence. Experimental studies of collision energy-dependent fragmentation efficiencies coupled with quantum chemical calculations indicated that the cleavage of ARRA and its side-chain oxidation products with oxygen atoms added followed a trend that doubly-protonated peptides fragment more easily than singly-protonated forms, while the oxidation product with the guanidine group deleted showed the opposite trend. By analyzing the charge distribution around the amide bonds, we found that the relative charge ratios between C and N atoms (QC/QN) in the amide bonds provided a reasonable explanation for peptide fragmentation efficiencies. An increase of the QC/QN value of the amide bond means that a peptide fragments more easily, and vice versa. The results described in this paper provide an experimental and calculation strategy for predicting peptide fragmentation efficiency.Graphical AbstractThe relative charge ratio between C and N atoms in amide bonds provides a reasonable explanation for peptide fragmentation efficiency.Download high-res image (163KB)Download full-size image
Co-reporter:Yue Teng, Fanying Ji, Chao Li, Zehua Yu, Rutao Liu
Journal of Luminescence (December 2011) Volume 131(Issue 12) pp:2661-2667
Publication Date(Web):1 December 2011
DOI:10.1016/j.jlumin.2011.07.005
4-Aminoantipyrine (AAP) is scarcely administered as a kind of analgesic drug because of the side effect. The residue of AAP in the environment poses a potential threat to human health. To evaluate the toxicity of AAP at the protein level, the effects of AAP on lysozyme were investigated using spectroscopic and molecular docking methods. Addition of AAP effectively quenched the intrinsic fluorescence of lysozyme. Static quenching of lysozyme by AAP revealed the formation of complex. After the inner filter effect was eliminated, the number of binding sites, the binding constant and the thermodynamic parameters were measured, and indicated that AAP can spontaneously bind with lysozyme through hydrophobic interactions with one binding site. Molecular docking results revealed that AAP bound into the enzyme active site and interacted with the Trp 62 and Trp 63 residues of lysozyme, which illustrated that the lysozyme activity was inhibited by AAP, in accordance with the results of the lysozyme activity experiment. Furthermore, the binding of AAP can result in demonstrable change of the conformation of lysozyme. This work is helpful for clarifying the molecular toxic mechanism of AAP in vivo.Highlights► This work established the binding mode of AAP with lysozyme in molecular level. ► Mechanism was explored by multiple spectroscopic and molecular docking methods. ► AAP can inhibit lysozyme activity and induce conformational changes in lysozyme.
Co-reporter:Jing Wang, Jinhu Wang, Wei Song, Xinping Yang, Wansong Zong and Rutao Liu
Physical Chemistry Chemical Physics 2016 - vol. 18(Issue 5) pp:NaN3544-3544
Publication Date(Web):2016/01/11
DOI:10.1039/C5CP06100H
Cadmium adversely affects the biological function of the liver. Transferrin might be involved in the detoxification system of cadmium. However, owing to the lack of investigation of the molecular mechanism of cadmium conjugating to transferrin, the role of transferrin in cadmium detoxification in the liver and how transferrin undergoes conformational and functional changes upon cadmium binding are not clear. In this article, we demonstrated the potential role of transferrin in the protection of the mouse primary hepatocytes against cadmium toxicity. After the incubation of hepatocytes with 10 and 100 μM CdCl2, pretreatment with transferrin significantly attenuated the reduction of cell viability in comparison with the samples treated with CdCl2 alone. Furthermore, a detailed molecular mechanism investigation of the interaction of CdCl2 with transferrin was reported using biophysical methods. Multi-spectroscopic measurements showed that CdCl2 formed complexes with transferrin and caused structural and conformational changes of transferrin. Isothermal titration calorimetry measurements revealed that transferrin has two classes of binding sites with different binding constants for CdCl2 binding. Hydrophobic forces and electrostatic forces are the major driving forces of the interaction. Preferred specific binding sites on transferrin were identified by dialysis experiments, molecular docking studies and molecular dynamics simulations. Upon low CdCl2 concentration exposure, no content of iron was released from transferrin because CdCl2 preferentially binds to the surface of transferrin molecules. Upon higher CdCl2 concentration exposure, the release of iron content from transferrin was observed due to the interaction of CdCl2 with the key residues around iron binding sites.
![[1,1'-Biphenyl]-3,3'-dipropanoicacid, a3,a3'-diamino-6,6'-dihydroxy-](http://img.cochemist.com/ccimg/1000/980-21-2.png)
![[1,1'-Biphenyl]-3,3'-dipropanoicacid, a3,a3'-diamino-6,6'-dihydroxy-](http://img.cochemist.com/ccimg/1000/980-21-2_b.png)
