Co-reporter:Xian Kong;Jianzhong Wu;Diannan Lu
Langmuir April 19, 2016 Volume 32(Issue 15) pp:3785-3793
Publication Date(Web):2017-2-22
DOI:10.1021/acs.langmuir.6b00043
Supported lipid bilayers (SLBs) are able to accommodate membrane proteins useful for diverse biomimetic applications. Although liposome spreading represents a common procedure for preparation of SLBs, the underlying mechanism is not yet fully understood, particularly from a molecular perspective. The present study examines the effects of the substrate charge on unilamellar liposome spreading on the basis of molecular dynamics simulations for a coarse-grained model of the solvent and lipid molecules. Liposome transformation into a lipid bilayer of different microscopic structures suggests three types of kinetic pathways depending on the substrate charge density, that is, top-receding, parachute, and parachute with wormholes. Each pathway leads to a unique distribution of the lipid molecules and thereby distinctive properties of SLBs. An increase of the substrate charge density results in a magnified asymmetry of the SLBs in terms of the ratio of charged lipids, parallel surface movements, and the distribution of lipid molecules. While the lipid mobility in the proximal layer is strongly correlated with the substrate potential, the dynamics of lipid molecules in the distal monolayer is similar to that of a freestanding lipid bilayer. For liposome spreading on a highly charged surface, wormhole formation promotes lipid exchange between the SLB monolayers thus reduces the asymmetry on the number density of lipid molecules, the lipid order parameter, and the monolayer thickness. The simulation results reveal the important regulatory role of electrostatic interactions on liposome spreading and the properties of SLBs.
Co-reporter:Weina XuYou Yong, Zheyu Wang, Guoqiang Jiang, Jianzhong Wu, Zheng Liu
ACS Sustainable Chemistry & Engineering 2017 Volume 5(Issue 1) pp:
Publication Date(Web):December 16, 2016
DOI:10.1021/acssuschemeng.6b02705
A facile method of immobilizing enzymes on activated carbon (AC) is proposed, in which the first step is to coat AC with Concanavalin A (ConA), followed by the adsorption of enzymes. Two model glycoenzymes, horseradish peroxidase and laccase, were immobilized on the ConA adsorbed AC through the tightly specific recognition of ConA to their glycosidic moieties, as confirmed by laser confocal microscopy. The coating layer of ConA reduced the deactivation of enzymes and prevented the leakage of enzymes, as indicated by the significantly improved yield of enzymatic activity. The immobilized enzymes on ConA coated AC appeared an improved stability against pH and temperature, offering an expanded operation “window” for growing applications of the enzymatic catalysis. Finally, the integration of the adsorption capacity of AC and the chemical degradation of laccase promises the efficient removal of aqueous phenol. The improved recovery of enzyme activity, enhanced stability as well as the combination of substrate adsorption and enzymatic reaction make ConA coated AC appealing for various enzymatic catalysis.Keywords: Activated carbon; Concanavalin A; Enzyme immobilization; Horseradish peroxidase; Laccase; Phenol removal;
Co-reporter:Gong Chen;Xian Kong;Diannan Lu;Jianzhong Wu
Physical Chemistry Chemical Physics 2017 vol. 19(Issue 18) pp:11690-11697
Publication Date(Web):2017/05/10
DOI:10.1039/C7CP00887B
Molecular dynamics (MD) simulations, in combination with the Markov-state model (MSM), were applied to probe CO2 diffusion from an aqueous solution into the active site of human carbonic anhydrase II (hCA-II), an enzyme useful for enhanced CO2 capture and utilization. The diffusion process in the hydrophobic pocket of hCA-II was illustrated in terms of a two-dimensional free-energy landscape. We found that CO2 diffusion in hCA-II is a rate-limiting step in the CO2 diffusion-binding-reaction process. The equilibrium distribution of CO2 shows its preferential accumulation within a hydrophobic domain in the protein core region. An analysis of the committors and reactive fluxes indicates that the main pathway for CO2 diffusion into the active site of hCA-II is through a binding pocket where residue Gln136 contributes to the maximal flux. The simulation results offer a new perspective on the CO2 hydration kinetics and useful insights toward the development of novel biochemical processes for more efficient CO2 sequestration and utilization.
Co-reporter:Yifei Zhang, You Yong, Jun Ge, and Zheng Liu
ACS Catalysis 2016 Volume 6(Issue 6) pp:3789
Publication Date(Web):May 6, 2016
DOI:10.1021/acscatal.6b01047
A method based on biological recognition was proposed to prepare cross-linked multiple glycoenzyme aggregates. With Concanavalin A (ConA) as a molecular glue, horseradish peroxidase (HRP) and glucose oxidase (GOx) were agglutinated and then cross-linked by glutaraldehyde, forming a GOx-ConA-HRP catalyst. The affinity of ConA to glucose enhanced the uptake of the substrate, reducing the Km of cross-linked GOx-ConA-HRP aggregates for glucose from 51 mM to 8.8 mM. The colocalization and clustering of cascade enzymes at nanoscale facilitated the intermediate consumption. These effects significantly improved the catalytic performance of the GOx-HRP cascade with a 1.5-fold increased specificity constant. The use of ConA as a molecular glue provides a facile way to construct a multienzyme catalyst with enhanced stability and activity.Keywords: Concanavalin A; enhanced activity; glycoenzyme aggregates; multienzyme systems; substrate affinity
Co-reporter:Oda Yuki;Yifei Zhang;Jun Ge
Catalysis Letters 2016 Volume 146( Issue 6) pp:1073-1078
Publication Date(Web):2016 June
DOI:10.1007/s10562-016-1726-5
In the present study, Candida antarctica lipase B (CALB) was conjugated with Pluronic polymers, a family of amphiphilic triblock copolymers made of a central hydrophobic polypropylene oxide block connected to two hydrophilic poly(ethylene oxide) side blocks, and tested for the chemo-enzymatic epoxidation of fatty acids conducted in organic media. The effects of operation parameters including the type of Pluronic polymers and solvents on the epoxidation of different plant oil substrates were examined, in which Pluronic F127 and toluene appeared the best polymer and solvent, respectively. At optimized conditions, an yield of 97, 75, 67 % for epoxidized oleic acid, linoleic acid, linolenic acid, respectively, was obtained using Pluronic F127 conjugated CALB. This study suggested that CALB-Pluronic conjugate is an appealing enzyme catalyst for the epoxidation in organic media.
Co-reporter:Yifei Zhang, Jun Ge, and Zheng Liu
ACS Catalysis 2015 Volume 5(Issue 8) pp:4503
Publication Date(Web):June 16, 2015
DOI:10.1021/acscatal.5b00996
Re-engineering enzymes with high activities in the given environments different from the physiological one has been constantly pursued for application of enzymatic catalysis in industrial biocatalytic processes, pharmaceutical industry, biosensing, etc. Re-engineering enzyme catalysts by chemical approaches, including immobilization and chemical modification, represents a simple but effective route. The unusual phenomenon that immobilized or chemically modified enzymes display higher activities than native enzymes has been observed in both single- and multiple-enzyme systems. Recent achievements in enhancing enzymatic activities in both single-and multiple-enzyme systems by chemical approaches are summarized in this review. We propose that these enhanced enzymatic activities can be attributed to the well-designed specific interactions between immobilization carriers (or chemical modifiers) and enzymes, substrates, or reaction media. In addition to this mechanism, which is applicable for both single- and multiple-enzyme systems, other important factors responsible for enhanced activities of multiple-enzyme systems, including substrate channeling, kinetic matching, and an ordered spatial distribution of enzymes, are also discussed. Understanding the origin of enhanced activity in enzymatic catalysis may provide new insights and inspiration to design efficient enzyme catalysts for practical applications.Keywords: chemically modified enzyme; enhanced enzymatic activity; enzymatic catalysis; immobilized enzyme; multienzyme system
Co-reporter:Gong Chen, Xian Kong, Jingying Zhu, Diannan Lu and Zheng Liu
Physical Chemistry Chemical Physics 2015 vol. 17(Issue 16) pp:10708-10714
Publication Date(Web):12 Mar 2015
DOI:10.1039/C5CP00418G
While the conjugation of enzymes with ABA copolymers has resulted in increased enzymatic activities in organic solvents, by several orders of magnitude, the underpinning mechanism has not been fully uncovered, particularly at the molecular level. In the present work, a coarse-grained molecular dynamics simulation of cytochrome c (Cyt c) conjugated with a PEO–PPO–PEO block copolymer (ABA) in toluene was simulated with Cyt c as a control. It is shown that the hydrophilic segments (PEO) of the conjugated block copolymer molecules tend to entangle around the hydrophilic patch of Cyt c, while the hydrophobic segments (PPO) extend into the toluene. At a lower temperature, the PEO tails tend to form a hairpin structure outside the conjugated protein, whereas the Cyt c–ABA conjugates tend to form larger aggregates. At a higher temperature, however, the PEO tails tend to adsorb onto the hydrophilic protein surface, thus improving the suspension of the Cyt c–ABA conjugates and, consequently, the contact with the substrate. Moreover, the temperature increase drives the conformational transition of the active site of Cyt c–ABA from an “inactive state” to an “activated state” and thus results in an enhanced activity. To validate the above simulations, Cyt c was conjugated to F127, an extensively used ABA copolymer. By elevating the temperature, a decrease in the average size of the Cyt c–F127 conjugates along with a great increase in the apparent activity in toluene was observed, as can be predicted from the molecular dynamics simulation. The above mentioned molecular simulations offer a molecular insight into the temperature-responsive behaviour of protein–ABA copolymers, which is helpful for the design and application of enzyme–polymer conjugates for industrial biocatalysis.
Co-reporter:Jipeng Li, Yiyun Ouyang, Xian Kong, Jingying Zhu, Diannan Lu and Zheng Liu
RSC Advances 2015 vol. 5(Issue 83) pp:68227-68233
Publication Date(Web):28 Jul 2015
DOI:10.1039/C5RA10965E
The capability of silencing genes makes small interfering RNA (siRNA) appealing for curing fatal diseases such as cancer and viral infections. In the present work, we chose a novel amphiphilic polymer, PMAL (poly(maleic anhydride-alt-1-decene) substituted with 3-(dimethylamino) propylamine), as the siRNA carrier, and conducted steered molecular dynamics simulations, together with traditional molecular dynamics simulations, to explore how PMAL facilitates the delivery of siRNA. It was shown that the use of PMAL reduced the energy barrier for siRNA to penetrate lipid bilayer membranes, as confirmed by the experimental work. The simulation of the transmembrane process revealed that PMAL can punch a hole in the lipid bilayer and form a channel for siRNA delivery. Monitoring of the structural transition further showed the targeting of siRNA through the attachment of PMAL encapsulating siRNA to a lipid membrane. The delivery of siRNA was facilitated by the hydrophobic interaction between PMAL and the lipid membrane, which favored the dissociation of the siRNA–PMAL complex. The above simulation established a molecular insight of the interaction between siRNA and PMAL and was helpful for the design and applications of new carriers for siRNA delivery.
Co-reporter:Zongjun Li;Yifei Zhang;Diannan Lu
Journal of Applied Polymer Science 2015 Volume 132( Issue 39) pp:
Publication Date(Web):
DOI:10.1002/app.42596
ABSTRACT
A cationic block copolymer (mPEG-b-PMETAC) with uniform molecular weight was synthesized using atom transfer radical polymerization. Insulin was reversibly encapsulated by mPEG-b-PMETAC via electrostatic interaction. The secondary and tertiary structures of insulin during the assembly and delivery processes were monitored by circular dichroism and fluorescence spectrum. The effects of pH and salt concentration on encapsulation were examined, respectively. Enhanced stability of the encapsulated insulin against proteolysis by trypsin and chymotrypsin was demonstrated. Insulin can be encapsulated and delivered from an mPEG-b-PMETAC assembly by tuning the pH and ionic strength, which determines the electrostatic interaction between insulin and the polymer. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015, 132, 42596.
Co-reporter:Liwei Ren;Yushen Wang;Jun Ge;Diannan Lu
Macromolecular Chemistry and Physics 2015 Volume 216( Issue 6) pp:636-640
Publication Date(Web):
DOI:10.1002/macp.201400550
Co-reporter:Yijie Dong, Zhe Lang, Xian Kong, Diannan Lu and Zheng Liu
Environmental Science: Nano 2015 vol. 17(Issue 4) pp:763-774
Publication Date(Web):10 Feb 2015
DOI:10.1039/C4EM00428K
Biostimulation, which employs nutrients to enhance the proliferation of indigenous microorganisms and therefore the degradation of contaminants, is an effective tool for treatment of oil-contaminated soil. However, the evolution of microbial ecology, which responds directly to stimulation procedures and intrinsically determines the degradation of oil contaminants, has rarely been explored, particularly in the context of biostimulation. In this study, the effects of biostimulation procedures including the regulation of the C:N:P ratio, as well as application of surfactants and electron acceptors in the degradation of crude oil contaminants and the evolution of the microbial community were examined simultaneously to provide ecological insights into the biostimulation. The real-time PCR showed that biostimulation promoted the proliferation of bacteria, with Gammaproteobacteria showing the greatest increase. However, the proliferation of fungi was inhibited by the accumulation of the degradation products. The degradation of polar compounds of crude oil contaminants was characterized by negative-ion electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (negative-ion ESI FT-ICR MS), showing a biased increase in the relative abundance of naphthenic acids. Principal component analysis (PCA) showed that different species in oil sludge have different degradation rates during biostimulation. The addition of fertilizers with surfactants and electron acceptors profoundly stimulated the indigenous microorganisms with N1, O1 and O2 species as substrates while those with O3 and O4 species were little affected. An enriched abundance of alkB genes was observed during the degradation of saturated hydrocarbons. Monitoring the kinetics of the microbial community, functional genes and degradation offers a comprehensive view for the understanding and optimization of the biostimulation process.
Co-reporter:Rui Wang;Miao Hou;Yifei Zhang;Jun Ge
Catalysis Letters 2015 Volume 145( Issue 4) pp:995-999
Publication Date(Web):2015 April
DOI:10.1007/s10562-015-1493-8
We report the first example of enzymatic synthesis of lutein dipalmitate in organic solvents by using a commercial immobilized Candida antarctica lipase B (Novozyme 435) as the catalyst. At the optimized condition, by using 20 mg/mL of lutein, 100 mg/mL of vinyl palmitate as the substrates, and 20 mg/mL of Novozyme 435 as the catalyst, the enzymatic synthesis in toluene achieved a yield of lutein dipalmitate of 83 % after 8 h reaction at 60 °C. The stability of the enzyme was demonstrated by over 80 % conversions for ten batches.
Co-reporter:Jipeng Li;Xian Kong;Diannan Lu
Science Bulletin 2015 Volume 60( Issue 18) pp:1580-1586
Publication Date(Web):2015 September
DOI:10.1007/s11434-015-0888-7
While the preferential movement of water inside carbon nanotube is appealing for water purification, our understanding of the water transport mechanism through carbon nanotube (CNT)-based membrane is far from adequate. Here we conducted molecular dynamics simulations to study how the alignment of the CNTs in the membrane affects the water transport through the CNT membrane. It was shown that compared to the conventional CNT membrane where the alignment of CNTs was vertical to membrane surface, the “italicized CNT membrane” in which the contact angel between membrane surface and the CNT alignment is not 90° offered a higher transmembrane flux of water. The expanded exposure of more carbon atoms to water molecules reduced the energy barrier near the entrance of this italicized CNT membrane, compared to the vertical one. For water flows through the italicized CNT membrane, the Lennard-Jones interaction between water and nanotube as function of central path of the CNT changes from “U” to “V” pattern, which significantly lowers energy barrier for filling water into the CNT, favoring the water transport inside carbon nanotube. Above simulation indicates new opportunities for applying CNT in water purification or related fields in which water transport matters.
Co-reporter:Fengjiao Lyu, Yifei Zhang, Richard N. Zare, Jun Ge, and Zheng Liu
Nano Letters 2014 Volume 14(Issue 10) pp:5761-5765
Publication Date(Web):September 11, 2014
DOI:10.1021/nl5026419
Protein molecules were directly embedded in metal–organic frameworks (MOFs) by a coprecipitation method. The protein molecules majorly embedded on the surface region of MOFs display high biological activities. As a demonstration of the power of such materials, the resulting Cyt c embedded in ZIF-8 showed a 10-fold increase in peroxidase activity compared to free Cyt c in solution and thus gave convenient, fast, and highly sensitive detection of trace amounts of explosive organic peroxides in solution.
Co-reporter:Yifei Zhang, Fengjiao Lyu, Jun Ge and Zheng Liu
Chemical Communications 2014 vol. 50(Issue 85) pp:12919-12922
Publication Date(Web):28 Aug 2014
DOI:10.1039/C4CC06158F
A method using ink-jet printing for constructing multi-enzyme systems was proposed, in which a precisely defined enzyme ratio and two-dimensional distribution was obtained by the preset ‘color’ values. The applications of the print-on-paper multi-enzyme systems were exemplified by the detection of glucose and the design of an enzyme-enabled two-dimensional code.
Co-reporter:Xian Kong, Shanshan Qin, Diannan Lu and Zheng Liu
Physical Chemistry Chemical Physics 2014 vol. 16(Issue 18) pp:8434-8440
Publication Date(Web):18 Mar 2014
DOI:10.1039/C3CP55524K
While the surface tension of a cell membrane, or a plasma membrane, regulates cell functions, little is known about its effect on the conformational changes of the lipid bilayer and hence the resulting changes in the cell membrane. To obtain some insights into the phase transition of the lipid bilayer as a function of surface tension, we used a 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayer as a model lipid bilayer and aquaporin (AqpZ), a transmembrane channel protein for water, as a model embedded protein. A coarse-grained molecular dynamics simulation was applied to illustrate the phase transition behavior of the pure DPPC bilayer and aquaporin-embedded DPPC bilayer under different surface tensions. It was shown that an increased surface tension reduced the phase transition temperature of the DPPC bilayer. As for the DPPC bilayer in gel form, no significant changes occurred in the structure of the bilayer in response to the surface tension. Once in a liquid crystal state, both the structure and properties of the DPPC bilayer, such as area per lipid, lipid order parameters, bilayer thickness and lateral diffusion coefficients, were responsive to the magnitude of surface tension in a linear way. The presence of aquaporin attenuated the compact alignment of the lipid bilayer, hindered the parallel movement, and thus made the DPPC bilayer less sensitive to the surface tension.
Co-reporter:Rui Wang, Yifei Zhang, Jun Ge and Zheng Liu
RSC Advances 2014 vol. 4(Issue 76) pp:40301-40304
Publication Date(Web):14 Aug 2014
DOI:10.1039/C4RA06660J
A substrate or polyethyleneglycol (PEG) imprinted lipase nanogel displayed increased apparent activity in organic solvents by 2.5–4.7 folds, compared to native lipase. It enabled a one-step synthesis of chloramphenicol palmitate with a yield of ∼99% and purity of ∼99%, indicating that the imprinted lipase nanogel is an appealing catalyst in organic media.
Co-reporter:Xian Kong, Jian Jiang, Diannan Lu, Zheng Liu, and Jianzhong Wu
The Journal of Physical Chemistry Letters 2014 Volume 5(Issue 17) pp:3015-3020
Publication Date(Web):August 19, 2014
DOI:10.1021/jz5013802
Ion transport through nanochannels depends on various external driving forces as well as the structural and hydrodynamic inhomogeneity of the confined fluid inside of the pore. Conventional models of electrokinetic transport neglect the discrete nature of ionic species and electrostatic correlations important at the boundary and often lead to inconsistent predictions of the surface potential and the surface charge density. Here, we demonstrate that the electrokinetic phenomena can be successfully described by the classical density functional theory in conjunction with the Navier–Stokes equation for the fluid flow. The new theoretical procedure predicts ion conductivity in various pH-regulated nanochannels under different driving forces, in excellent agreement with experimental data.Keywords: density functional theory; electric double layer; electro-osmotic flow; ion transport; nanofluidics; Poisson−Boltzmann equation;
Co-reporter:Xiaoling Wu;Rui Wang;Yifei Zhang;Jun Ge
Catalysis Letters 2014 Volume 144( Issue 8) pp:1407-1410
Publication Date(Web):2014 August
DOI:10.1007/s10562-014-1289-2
Asymmetrical ammonolysis of (R)- and (S)-phenylglycine methyl ester was carried out by using a lipase (CALB)–polymer (Pluronic) nanoconjugate as the catalyst, displaying a 11-fold increased catalytic rate compared to the free CALB in tertiary butanol.
Co-reporter:Binbin Wu, Tian Lan, Diannan Lu and Zheng Liu
Environmental Science: Nano 2014 vol. 16(Issue 6) pp:1501-1509
Publication Date(Web):28 Feb 2014
DOI:10.1039/C3EM00731F
The changes in microbial ecology interpreted from taxonomic and functional genes and biological functions represented by urease and dehydrogenase activities were monitored in soil contaminated with different petroleum hydrocarbons including crude oil, diesel, n-hexadecane and poly-aromatic hydrocarbons (PAHs). It was shown that the presence of n-hexadecane stimulated the activity of indigenous microorganisms, especially alkane degrading bacteria, and led to over 20% degradation of n-hexadecane within one month. No obvious degradation of the other three types of petroleum hydrocarbons was observed. The stimulation effect was most marked in the soil spiked with a medium concentration (2500 mg kg−1 dry soil) of n-hexadecane. However, the presence of PAHs completely inhibited the previously-mentioned bioactivities of the soil. The content of PAH degrading bacteria, however, increased more than 10-fold, indicating the selection effect of PAHs on soil bacteria. The impacts of diesel and crude oil on the microbial ecology and biological functions varied significantly with their concentration. The disclosure of the ecological and enzymatic responses could be helpful in soil bioremediation.
Co-reporter:Rui Wang, Yifei Zhang, Jinhai Huang, Diannan Lu, Jun Ge and Zheng Liu
Green Chemistry 2013 vol. 15(Issue 5) pp:1155-1158
Publication Date(Web):19 Mar 2013
DOI:10.1039/C3GC40465J
Enzymatic catalysis with high enantio- and regio-selectivity, which is attractive for green synthesis of chemicals, often suffers from low activity in organic solvents utilized as reaction media. Here, we describe a ‘substrate-imprinted’ lipase nanogel that displays high activity in organic solvents. The first step was to encapsulate lipase into polyacrylamide nanogel by an aqueous in situ polymerization. Then the lipase nanogel was lyophilized in the presence of palmitic acid, a substrate of lipase, followed by extraction with petroleum ether to remove palmitic acid from the lyophilized lipase nanogel. The imprinting treatment increased the adsorption capacity of palmitic acid by 2.9-fold and the apparent activity by 2-fold in catalyzing the transesterification reaction between para-nitrophenyl palmitate and ethanol. The effects of solvent and temperature on the yield and selectivity of the enzymatic synthesis of chloramphenicol palmitate were examined, respectively. One-step synthesis of chloramphenicol palmitate with the imprinted lipase nanogel gave a yield of ∼99% and a purity of ∼99% within 12 hours at 20 °C, whereas the imprinted free lipase gave a yield below 60% in 20 hours. The high activity and selectivity make the substrate-imprinted enzyme nanogel an attractive catalyst for green synthesis of chemicals having complex structures.
Co-reporter:Yifei Zhang, Qile Chen, Jun Ge and Zheng Liu
Chemical Communications 2013 vol. 49(Issue 84) pp:9815-9817
Publication Date(Web):29 Aug 2013
DOI:10.1039/C3CC45837G
An enzyme-incorporated hydrogel made of alginate and polyacrylamide shows a linearly increased activity with the enlargement of surface area upon stretching.
Co-reporter:Jingying Zhu, Yifei Zhang, Diannan Lu, Richard N. Zare, Jun Ge and Zheng Liu
Chemical Communications 2013 vol. 49(Issue 54) pp:6090-6092
Publication Date(Web):22 May 2013
DOI:10.1039/C3CC42493F
A general approach for preparing enzyme–polymer nanoconjugates that respond to temperature in organic media is presented. These nanoconjugates readily dissolve in organic solvents for homogenous catalysis at 40 °C and showed greatly enhanced apparent catalytic activities. The recovery of the soluble enzyme–polymer nanoconjugates is accomplished by temperature-induced precipitation.
Co-reporter:Yifei Zhang, Kehang Han, Diannan Lu and Zheng Liu
Soft Matter 2013 vol. 9(Issue 36) pp:8723-8729
Publication Date(Web):18 Jul 2013
DOI:10.1039/C3SM50586C
We synthesized a novel hydrophilic, negatively charged block polymer composed of polyethylene glycol (PEG) and poly(methacrylic acid) (PMAA) using atom-transfer radical polymerization (ATRP). The encapsulation of a positively charged protein, represented by hen-egg white lysozyme, by mPEG-b-PMAA micelles was achieved using a pH or salt-concentration swing, as shown by both structural characterization using dynamic light scattering and transmission electron microscopy and an activity assay. All-atom molecular dynamics simulations showed that using an acidic pH gave a more compact polymer–micelle assembly than did using a basic pH. As a result, this compact structure had less solvent-accessible surface area (SASA), indicating that lysozyme was encapsulated by mPEG-b-PMAA and that the active site was shielded by the polymer. This made the active site less accessible to the substrate. These accounted for the low apparent activity at an acidic pH in our experiments. A neutral or basic pH intensified the electrostatic repulsive interaction, which prevented the formation of polymer–lysozyme complex. The molecular simulation indicated that encapsulation of lysozyme by the polymer micelles could be divided into two consecutive steps. The first step involved the attachment of the negatively charged polymer chain to the positively charged portion of lysozyme, driven by electrostatic attractive force. Then, the hydrophobic interaction between the polymer and lysozyme became dominant and led to a more compact assembly with a reduced energy state. These simulations agreed with our experimental observations and provided molecular insight helpful for the design, fabrication, and application of protein-incorporated polymer micelles.
Co-reporter:Zhixian Li, Yifei Zhang, Mengmeng Lin, Pingkai Ouyang, Jun Ge, and Zheng Liu
Organic Process Research & Development 2013 Volume 17(Issue 9) pp:1179-1182
Publication Date(Web):August 12, 2013
DOI:10.1021/op400135y
Chemical synthesis of clindamycin palmitate, a prodrug with taste greatly improved more than that of clindamycin, involves laborious steps of protection and deprotection to achieve the monoacylation only at 2-hydroxyl group of clindamycin and gives an overall yield below 50%. Here we report the first example of one-step synthesis of clindamycin palmitate with high regioselectivity using immobilized Candida antarctica lipase B (Novozym 435) as the catalyst. The lipase-catalyzed synthesis reached a conversion above 90% in 12 h using toluene as solvent and, moreover, a highly regioselective acylation at the 2-hydroxyl of clindamycin. The significantly improved conversion achieved at an excellent regioselectivity makes this enzymatic process attractive for the synthesis of clindamycin ester derivatives.
Co-reporter:Yifei Zhang, Yang Dai, Miao Hou, Tian Li, Jun Ge and Zheng Liu
RSC Advances 2013 vol. 3(Issue 45) pp:22963-22966
Publication Date(Web):03 Oct 2013
DOI:10.1039/C3RA44879G
A novel synthetic route for valrubicin, an anti-cancer drug, was proposed using a temperature-responsive lipase–Pluronic conjugate which gave an overall yield of 82% and a purity of 98% under mild conditions (50 °C, 6–8 h). The high activity, stability, selectivity and reusability of this new lipase catalyst hold great promise for practical applications.
Co-reporter:Mengmeng Lin, Diannan Lu, Jingying Zhu, Cheng Yang, Yifei Zhang and Zheng Liu
Chemical Communications 2012 vol. 48(Issue 27) pp:3315-3317
Publication Date(Web):13 Feb 2012
DOI:10.1039/C2CC30189J
A universal synthetic route for magnetic enzyme nanogels (MENGs) was proposed, based on electrostatic interaction driven assembly and in situ polymerization from the surface of magnetic nanoparticles, to avoid chemical modification of proteins and hence structural and functional deterioration.
Co-reporter:Dandan Xu, Lige Tonggu, Xiaoping Bao, Diannan Lu and Zheng Liu
Soft Matter 2012 vol. 8(Issue 6) pp:2036-2042
Publication Date(Web):03 Jan 2012
DOI:10.1039/C1SM06853A
A lipase nanogel has been prepared by aqueous in situ polymerization, with initial acryloylation to introduce vinyl groups onto the lipase surface for subsequent polymerization with acrylamide monomers. Activation of lipase was observed using glycidyl methacrylate (GMA) as the acryloylation agent, leading to an increase of activity yield from 78 ± 4% (obtained using NAS for acryloylation) to 122 ± 14%. The acryloylation ratio was also improved, from 8 ± 5% to 20 ± 8%, which favored the subsequent polymerization. In addition, the overall activity yield of the lipase nanogel was increased from 44 ± 3% to 105 ± 2%. The lipase nanogel prepared with GMA for acryloylation displayed a lower Km and a higher kcat in comparison to its native counterpart, as well as the nanogel obtained using NAS for acryloylation. The half-life of the lipase nanogel at 60 °C was extended from 1.63 h (using NAS for acryloylation) to 5.61 h, while the half-life of the native lipase was 1.41 h. Molecular dynamics simulations and experiments further suggested that the activation effect was due in part to interactions between the lipase and the GMA, which reinforced the ‘open’ configuration of the lipase, thereby facilitating mass transport to and from the modified lipase in the free and encapsulated forms. The above mentioned activation effect of the enzyme nanogel illustrated the power of chemical modification by tailored design of the nanostructure of the enzyme catalyst, potentially expanding the applications of enzyme catalysis.
Co-reporter:Liangdong Zou;Diannan Lu
Applied Biochemistry and Biotechnology 2012 Volume 168( Issue 7) pp:1976-1988
Publication Date(Web):2012 December
DOI:10.1007/s12010-012-9911-5
2,4,6-Trinitrotoluene (TNT), an extensively used and versatile explosive, is harmful in soil and water. In the present study, four bacterial strains capable of degrading TNT have been isolated from contaminated sites and named as Thu-A, Thu-B, Thu-C, and Thu-Z. Thu-Z, which gave the highest degradation efficiency compared to the others, was assigned to the genus Pantoea according to its 16S rRNA gene. Similarities in both biochemical properties and morphology suggested that Thu-Z was a Pantoea sp. strain. Thu-Z was proved to be capable of using TNT as a sole nitrogen source by cleaving NO2 from the nitroaromatic ring by direct aromatic ring reduction. Under nitrogen-limited conditions, 96.6 % N of TNT was consumed by Thu-Z for growth, which was determined in terms of NaNO2. Trace nitro reduction metabolites such as 2,4-diamino-6-nitrotoluene (24Dam) and 2,6-diamino-4-nitrotoluene (26Dam) were identified in the presence of (NH4)2SO4. On the other hand, 4,4′,6,6′-tetranitro-2,2′-azoxytoluene (22Azo) and 2,2′,6,6′-tetranitro-4,4′-azoxytoluene (44Azo) were detected in the absence of (NH4)2SO4. These indicated the existence of a dual pathway for Thu-Z, while the direct aromatic ring reduction was predominant. Addition of a nitrogen source ((NH4)2SO4) after inoculation stimulated the growth of Thu-Z and accelerated TNT degradation.
Co-reporter:Jun Ge;Cheng Yang;Jingying Zhu;Diannan Lu
Topics in Catalysis 2012 Volume 55( Issue 16-18) pp:1070-1080
Publication Date(Web):2012 November
DOI:10.1007/s11244-012-9906-z
In this review, we emphasized the importance of enzymatic processes in organic media, summarized recent advances of nanobiocatalysts with high activities in organic media, and proposed three general principles for designing nanobiocatalysis therein: facilitated substrate transport, retention of protein structure, and highly dispersed catalyst forms.
Co-reporter:Diannan Lu, Cheng Yang, and Zheng Liu
The Journal of Physical Chemistry B 2012 Volume 116(Issue 1) pp:390-400
Publication Date(Web):November 27, 2011
DOI:10.1021/jp203926r
Glycosylation is one of the most common post-translational modifications in the biosynthesis of protein, but its effect on the protein conformational transitions underpinning folding and stabilization is poorly understood. In this study, we present a coarse-grained off-lattice 46-β barrel model protein glycosylated by glycans with different hydrophobicity and glycosylation sites to examine the effect of glycans on protein folding and stabilization using a Langevin dynamics simulation, in which an H term was proposed as the index of the hydrophobicity of glycan. Compared with its native counterpart, introducing glycans of suitable hydrophobicity (0.1 < H < 0.4) at flexible peptide residues of this model protein not only facilitated folding of the protein but also increased its conformation stability significantly. On the contrary, when glycans were introduced at the restricted peptide residues of the protein, only those hydrophilic (H = 0) or very weak hydrophobic (H < 0.2) ones contributed slightly to protein stability but hindered protein folding due to increased free energy barriers. The glycosylated protein retained the two-step folding mechanism in terms of hydrophobic collapse and structural rearrangement. Glycan chains located in a suitable site with an appropriate hydrophobicity facilitated both collapse and rearrangement, whereas others, though accelerating collapse, hindered rearrangement. In addition to entropy effects, that is, narrowing the space of the conformations of the unfolded state, the presence of glycans with suitable hydrophobicity at suitable glycosylation site strengthened the folded state via hydrophobic interaction, that is, the enthalpy effect. The simulations have shown both the stabilization and the destabilization effects of glycosylation, as experimentally reported in the literature, and provided molecular insight into glycosylated proteins. The understanding of the effects of glycans with different hydrophobicities on the folding and stability of protein, as attempted by the present work, is helpful not only to explain the stabilization and destabilization effect of real glycoproteins but also to design protein–polymer conjugates for biotechnological purposes.
Co-reporter:Binbin Zhu, Diannan Lu, Jun Ge, Zheng Liu
Acta Biomaterialia 2011 Volume 7(Issue 5) pp:2131-2138
Publication Date(Web):May 2011
DOI:10.1016/j.actbio.2011.01.033
Abstract
In situ aqueous activators generated by electron transfer for atom transfer radical polymerization (AGET ATRP) in air, using an enzyme as a macroinitiator, has been proposed to prepare uniform polymer–protein conjugates with improved stability under adverse conditions. In the first step, an initiator, 2-bromoisobutyryl bromide (BIB), was grafted onto the protein surface by reaction with the amino groups. The second step was in situ AGET ATRP polymerization in air using CuBr2/1,1,4,7,7-pentamethyldiethylenetriamine as a catalyst and ascorbic acid as a reducing agent. The effectiveness of this method has been demonstrated using horseradish peroxidase (HRP) as a model protein and acrylamide as the monomer, which yielded HRP–polyacrylamide conjugate with a mean particle size of about 20–30 nm. The grafting of BIB onto HRP and the subsequent polymerization yielding a polyacrylamide chain were confirmed by nuclear magnetic resonance and matrix-assisted laser desorption ionization time-of-flight spectrometry analysis. The size of the conjugate was shown to be a function of monomer loading and reaction time. The HRP conjugates yielded essentially retained the catalytic behavior of HRP in free form, as shown by Km and Vmax values, but exhibited significantly enhanced thermal stability against high temperature and trypsin digestion. The use of protein as the macroinitiator prevented the formation of copolymer and thus facilitated purification of the protein conjugate. The uniform size indicates a well-defined composition of protein and polymer, which is essential for applications that request a precise control of the dosage of enzyme activity.
Co-reporter:Jun Ge;Diannan Lu;Cheng Yang
Macromolecular Rapid Communications 2011 Volume 32( Issue 6) pp:546-550
Publication Date(Web):
DOI:10.1002/marc.201000746
Co-reporter:Cheng Yang, Diannan Lu, and Zheng Liu
Biochemistry 2011 Volume 50(Issue 13) pp:
Publication Date(Web):February 18, 2011
DOI:10.1021/bi101926u
While the effectiveness of PEGylation in enhancing the stability and potency of protein pharmaceuticals has been validated for years, the underlying mechanism remains poorly understood, particularly at the molecular level. A molecular dynamics simulation was developed using an annealing procedure that allowed an all-atom level examination of the interaction between PEG polymers of different chain lengths and a conjugated protein represented by insulin. It was shown that PEG became entangled around the protein surface through hydrophobic interaction and concurrently formed hydrogen bonds with the surrounding water molecules. In addition to enhancing its structural stability, as indicated by the root-mean-square difference (rmsd) and secondary structure analyses, conjugation increased the size of the protein drug while decreasing the solvent accessible surface area of the protein. All these thus led to prolonged circulation life despite kidney filtration, proteolysis, and immunogenic side effects, as experimentally demonstrated elsewhere. Moreover, the simulation results indicated that an optimal chain length exists that would maximize drug potency underpinned by the parameters mentioned above. The simulation provided molecular insight into the interaction between PEG and the conjugated protein at the all-atom level and offered a tool that would allow for the design of PEGylated protein pharmaceuticals for given applications.
Co-reporter:Zhixia Liu, Diannan Lu, Ling Yin, Jianmin Li, Yuanchen Cui, Wei Chen, and Zheng Liu
The Journal of Physical Chemistry B 2011 Volume 115(Issue 28) pp:8875-8882
Publication Date(Web):June 23, 2011
DOI:10.1021/jp108941f
Urate oxidase (UOX, EC 1.7.3.3) is effective for the treatment of gout and hyperuricaemia associated with tumor lysis syndrome. The inherent poor stability of UOX to temperature, proteolysis, and acidic environments is known to limit its efficacy. Herein, we encapsulated UOX into spherical and porous nanogels with diameters of 20–40 nm via a two-step in situ polymerization in the presence of oxonic acid potassium salt, an inhibitor of UOX. The UOX nanogel retained 70% of the initial activity but showed an expanded pH spectrum from pH 6–10 to 3–10 and an extended half-life at 37 °C from 5 min to 3 h. The enhanced pH stability, thermal stability, and enzyme resistance of the UOX nanogels were also confirmed by using fluorescence spectroscopy and enzymatic digestion. A molecular dynamics simulation was performed as a way to probe the mechanism underlying the formation of UOX nanogels as well as the strengthened stability against harsh conditions. It was shown that the encapsulation into the polyacrylamide network reinforced the intersubunit hydrogen bonding, shielded the hydrolytic reaction site, and thus protected the tertiary and quaternary structure of UOX. The UOX nanogel with enhanced stability provided a stable enzyme model that enables the exploration of UOX in the diagnosis and therapy of disorders associated with altered purine metabolism.
Co-reporter:Zuojun Wu, Liangdong Zou, Diannan Lu and Zheng Liu
Environmental Science: Nano 2011 vol. 13(Issue 10) pp:2904-2913
Publication Date(Web):08 Sep 2011
DOI:10.1039/C0EM00761G
Soil microbial ecosystems are responsive to environmental changes that underpin the biological functions of the soil. The present study was conducted to profile variations in the microbial ecological system of remediated soil (R) and petroleum contaminated soil (P) based on comparisons with soil that had not been contaminated (N), using a cloning library of taxonomic genes (16S rRNA gene for bacteria and 18S rRNA gene for eukaryotes) and functional genes (nifH, amoA and narG). The results showed that N and R had a similar distribution in both the taxonomic genes and functional genes for bacteria and eukaryotes, which were dominated by Proteobacteria and Arthropoda, respectively. Phylogenetic analysis based on the nifHgene showed that the sequences from the three soils were clustered into six taxonomic groups, Actinobacteridae, and Alpha-, Beta-, Gamma- and Delta-proteobacteria, as well as an unclassified group. Evaluation of the amoAgene revealed that all sequences derived from the three samples belonged to Betaproteobacteria. The R and N soil had similar Shannon–Wiener diversity index (H′) values, both of which were significantly higher than that of the P soil. The most abundant bacterial phylotype identified in the N and R soils were the same and were related to an uncultured bacterial clone (GAN-SB17, FN423475). None of the narGgenes were found in the P soil. Similar results in terms of distribution, composition and the related index were obtained for nifH and amoA. These parameters may comprise a biological ecology index that may be applied to aid the design, implementation and evaluation of soil bioremediation.
Co-reporter:Zuojun Wu;Hongjun Dong;Liangdong Zou
Applied Biochemistry and Biotechnology 2011 Volume 164( Issue 7) pp:1071-1082
Publication Date(Web):2011 August
DOI:10.1007/s12010-011-9195-1
The bioaugmentation of petroleum-contaminated soil using Enterobacter cloacae was profiled from the evolution of microbial community, soil dehydrogenase activity, to the degradation of petroleum contaminants. The seeding and proliferation of inoculant and the consequential microbial community were monitored by denaturing gradient gel electrophoresis analysis of the amplification of V3 zone of 16S rDNA. Degradation process kinetics was characterized by the degradation ratio of nC17 to nC18. The dehydrogenase activity was also determined during the degradation process. An abrupt change in the microbial community after inoculation was illustrated as well as successive changes in response to degradation of the petroleum contaminants. Seeding with E. cloacae stimulated the growth of other degrading stains such as Pseudomonas sp. and Rhodothermus sp. The application of wheat straw as a representative lignin waste, at 5% (w/w), induced an increase in the total dehydrogenase activity from 0.50 to 0.79, an increase in the microbial content of 130% for bacteria and 84% for fungi, and an increase of the overall degradation ratio from 44% to 56% after 56 days of treatment. The above mentioned results have provided a microbial ecological insight being essential for the design and implementation of bioaugmentation processes.
Co-reporter:Zhixia Liu, Diannan Lu, Jianmin Li, Wei Chen and Zheng Liu
Physical Chemistry Chemical Physics 2009 vol. 11(Issue 2) pp:333-340
Publication Date(Web):24 Nov 2008
DOI:10.1039/B811496J
The aim of this study was to obtain molecular insight into the deactivation of recombinant urate oxidase (uricase, UOX, EC 1.7.3.3) (rUOX) from Aspergillus flavus. The enzyme is a tunnel-shaped homotetramer and has important clinical applications. By means of molecular dynamics simulations, multidimensional structural characterization and enzyme activity assays, we concluded that the thermal deactivation of UOX at neutral pH was associated with the loss of intersubunit hydrogen (H) bonds. This mechanism could also explain the deactivation of dilute aqueous UOX. Thermal deactivation of aqueous UOX due to dissociation of its subunits was ruled out. Displacement of H2O from the surface of UOX by less polar solvents such as methanol and dimethyl sulfoxide (DMSO) was proposed as an approach for strengthening intersubunit H bonds and consequently UOX stability. The effectiveness of this method was validated by both in silico and in vitro experiments. The results mentioned above provide insights for improving the stability of UOX and extending its applications. They may also be helpful for understanding the properties of other multimeric proteins.
Co-reporter:Lin Zhang, Diannan Lu, Zheng Liu
Journal of Chromatography A 2009 Volume 1216(Issue 12) pp:2483-2490
Publication Date(Web):20 March 2009
DOI:10.1016/j.chroma.2009.01.038
Conformational transitions of a protein in hydrophobic interaction based chromatography, including hydrophobic interaction chromatography (HIC) and reversed-phase liquid chromatography (RPLC), and their impact on the separation process and performance were probed by molecular dynamics simulation of a 46-bead β-barrel coarse-grained model protein in a confined pore, which represents the porous adsorbent. The transition of the adsorbed protein from the native conformation to an unfolded one occurred as a result of strong hydrophobic interactions with the pore surface, which reduced the formation of protein aggregates. The conformational transition was also displayed in the simulation once an elution buffer characterized by weaker hydrophobicity was introduced to strip protein from pore surface. The discharged proteins that underwent conformational transition were prone to aggregation; thus, an unsatisfactory yield of the native protein was obtained. An orthogonal experiment revealed that in addition to the strengths of the protein–protein and protein–adsorbent hydrophobic interactions, the elution time required to reduce the above-mentioned interactions also determined the yield of native protein by HIC and RPLC. Stepwise elution, characterized by sequential reduction of the hydrophobic interactions between the protein and adsorbent, was presented as a dynamic strategy for tuning conformational transitions to favor the native conformation and reduce the formation of protein aggregates during the elution process. The yield of the native protein obtained by this dynamic operation strategy was higher than that obtained by steady-state elution. The simulation study qualitatively reproduced the experimental observations and provided molecular insight that would be helpful for designing and optimizing HIC and RPLC separation of proteins.
Co-reporter:Jun Ge, Diannan Lu, Jun Wang and Zheng Liu
Biomacromolecules 2009 Volume 10(Issue 6) pp:
Publication Date(Web):April 10, 2009
DOI:10.1021/bm900205r
The present work showed that Candida rugosa lipase, which is inactive in anhydrous dimethyl sulfoxide (DMSO), has been granted its original catalytic activity and greatly enhanced stability when encapsulated into a polyacrylamide nanogel. The molecular simulation and structural analysis suggested that the polyacrylamide nanogel shielded the extraction of essential water and maintained the native configuration of encapsulated lipase in anhydrous DMSO at an elevated temperature. The electron and fluorescence microscopy showed that the lipase nanogel would be well dispersed in anhydrous DMSO where its native counterpart aggregated. The encapsulated lipase behaved as a stable catalyst for transesterification between dextran and vinyl decanoate in anhydrous DMSO at 60 °C for 240 h and yielded a dextran-based polymeric surfactant with regioselectivity toward the C-2 hydroxyl group in the glucopyranosyl unit of dextran. All these indicated a high potential of enzyme nanogel for nonaqueous biocatalysis.
Co-reporter:Diannan Lu and Zheng Liu
The Journal of Physical Chemistry B 2008 Volume 112(Issue 47) pp:15127-15133
Publication Date(Web):October 28, 2008
DOI:10.1021/jp804649g
One challenge in protein refolding is to dissociate the non-native disulfide bonds and promote the formation of native ones. In this study, we present a coarse-grained off-lattice model protein containing disulfide bonds and simulate disulfide bond shuffling during the folding of this model protein. Introduction of disulfide bonds in the model protein led to enhanced conformational stability but reduced foldability in comparison to counterpart protein without disulfide bonds. The folding trajectory suggested that the model protein retained the two-step folding mechanism in terms of hydrophobic collapse and structural rearrangement. The disulfide bonds located in the hydrophobic core were formed before the collapsing step, while the bonds located on the protein surface were formed during the rearrangement step. While a reductive environment at the initial stage of folding favored the formation of native disulfide bonds in the hydrophobic core, an oxidative environment at a later stage of folding was required for the formation of disulfide bonds at protein surface. Appling a dynamic redox environment, that is, one that changes from reductive to oxidative, intensified disulfide bond shuffling and thus resulted in improved recovery of the native conformation. The above-mentioned simulation was experimentally validated by refolding hen-egg lysozyme at different urea concentrations and oxidized glutathione/reduced glutathione (GSSG/GSH) ratios, and an optimal redox environment, in terms of the GSSG to GSH ratio, was identified. The implementation of a dynamic redox environment by tuning the GSSG/GSH ratio further improved the refolding yield of lysozyme, as predicted by molecular simulation.
Co-reporter:Jun Ge, Diannan Lu, Jun Wang, Ming Yan, Yunfeng Lu and Zheng Liu
The Journal of Physical Chemistry B 2008 Volume 112(Issue 45) pp:14319-14324
Publication Date(Web):October 21, 2008
DOI:10.1021/jp8053923
The assembly of a monomer around an enzyme as the essential step in the fabrication of enzyme nanogel by in situ polymerization was illustrated by molecular dynamics simulation and evidenced by a fluorescence resonance energy transfer spectrum, using lipase/acrylamide as a model system. The subsequent polymerization generated a hydrophilic gel network which not only strengthened the protein structural integrity via multipoint linkage but also increased the number of intramolecular H-bonds of the encapsulated protein, as suggested by the blue shift of the fluorescence spectrum of the encapsulated lipase. This greatly enhanced the stability of lipase at high temperature, as experimentally demonstrated. The exclusion of polar solvent molecules from the encapsulated enzyme, in contrast to the enrichment of water molecules, due to the presence of a hydrophilic gel network was displayed. This established a hydrophilic microenvironment for the encapsulated protein and thus gave the encapsulated protein an enhanced tolerance to the organic solvent, as experimentally observed in the present study and reported elsewhere. These results have given a molecular insight into the enzyme nanogel as well as its high potential as a robust enzyme model for an expended application spectrum of enzymatic catalysis.
Co-reporter:Wenguo Dong, Ming Yan, Zheng Liu, Guoshi Wu, Yanmei Li
Separation and Purification Technology 2007 Volume 53(Issue 2) pp:183-188
Publication Date(Web):25 February 2007
DOI:10.1016/j.seppur.2006.06.023
The influence of the solvent on the molecular recognition capability of molecularly imprinted polymers (MIPs) was studied. The energy difference (ΔE) of a molecule in vacuum and in a given solvent was calculated using density functional theory (DFT) at B3LYP/6-31+G** level, and used as a measure of the affinity of the solvent to the molecule. The ΔE of the template molecule, theophylline (THO), in three of the most extensively used solvents, chloroform, tetrahydrofuran (THF) and dimethyl sulfoxide (DMSO), was calculated, in which chloroform gave the smallest ΔE and DMSO gave the largest. The same order for ΔE was obtained for the monomer, methacrylic acid (MAA), in the three solvents. The calculated results indicate that chloroform is the most suitable solvent for the preparation of MIP for THO. DMSO, however, is not since it has high affinities to THO and MAA, and these inhibit the interaction between the THO and MAA. To examine the validity of the simulated results, MIPs for THO were synthesized in the three solvents and used for the adsorption of THO, respectively. The MIP synthesized in chloroform showed the maximum selectivity while that synthesized in DMSO showed the worst, in agreement with the above molecular simulation. H NMR spectroscopy showed that the H-bond between THO and MAA is the major force in molecular recognition, and this can be much affected by the solvent. The results above are of fundamental importance for the development of MIPs.
Co-reporter:Gang Yin, Jan-Christer Janson, Zheng Liu
Journal of Membrane Science 2000 Volume 178(1–2) pp:99-105
Publication Date(Web):15 September 2000
DOI:10.1016/S0376-7388(00)00484-1
A new method for determining the amount of adsorbed protein on membrane surface is developed on the basis of immunoassay using alkaline phosphotase as enzyme label and 4-nitrophenyl phosphate as substrate. Adsorption of human serum albumin (HSA) on HT Tuffryn, a kind of polysulfone microfiltration membrane, in the absence and presence of an electric field is investigated experimentally. It is shown that the adsorption of HSA on HT membrane surface is finished within 5 min and the amount of the adsorbed HSA decreases with the increase in pH but is not sensitive to salt concentration. Desorption of the adsorbed HSA from HT membrane surface can only be achieved with NaOH. Adding surfactants in HSA solution to prevent the adsorption is attempted and Tween 20 shows the best shielding function compared to polyethylene glycol 6000 and pluronic-F108. A critical concentration exists for Tween 20, below which the increase in the concentration of Tween 20 results in a corresponding reduction of adsorbed HSA. Introducing electric field into the adsorption leads to an enhanced adsorption of HSA, which appears to be a function of pH and the electric field strength.
Co-reporter:Zheng Liu;Shaohua Feng;Suhui Guo;Zhongyao Shen;Fuxin Ding;Naiju Yuan
Journal of Molecular Recognition 1998 Volume 11(Issue 1‐6) pp:151-156
Publication Date(Web):17 FEB 1999
DOI:10.1002/(SICI)1099-1352(199812)11:1/6<151::AID-JMR412>3.0.CO;2-6
A new method for preparative-scale separation of biomolecules, electrophoretic affinity chromatography (EAC), is proposed in this paper. Separation by EAC is carried out in a long and ribbon-like multicompartment electrolyser separated by membranes, in which the two central compartments are used for packing the gel matrix and for sample loading respectively. Next to the central compartments are the elution compartments and electrode compartments. The electric field is applied perpendicular to the fluid flow in the compartments. Adsorption and desorption steps may both be carried out in the presence of an electric field, which transports the target components into the gel compartment for adsorption and the impurities into the elution compartments for washing. After the adsorption step an elution solution is introduced and the product is released from the gel matrix and washed out. Separation of human serum albumin (HSA) from human serum gives HSA product of high purity, as demonstrated by isoelectric focusing analysis. The characteristics of electrophoretic binding of HSA on Blue Sepharose Fast Flow are examined. The preliminary results show that this new method has advantages in terms of high rate of mass transfer and ease of scaling up, which are of particular interest when large-scale separation of biomolecules is considered. Copyright © 1998 John Wiley & Sons, Ltd.
Co-reporter:Zheng Liu;Jin Wang;Jian Luo;Fuxin Ding;Naiju Yuan
Journal of Molecular Recognition 1998 Volume 11(Issue 1‐6) pp:149-150
Publication Date(Web):17 FEB 1999
DOI:10.1002/(SICI)1099-1352(199812)11:1/6<149::AID-JMR411>3.0.CO;2-N
Multichannel flow electrophoresis (MFE) is a newly developed method for continuous separation of biological products at a preparative scale, In this short survey, the application of MFE in the separation of proteins, enzymes and antibodies are overviewed. Copyright © 1998 John Wiley & Sons, Ltd.
Co-reporter:Mingliu Du, Diannan Lu, Zheng Liu
Journal of Molecular Catalysis B: Enzymatic (April 2013) Volume 88() pp:60-68
Publication Date(Web):1 April 2013
DOI:10.1016/j.molcatb.2012.11.017
A temperature-responsive lipase nanogel (denoted as CRL-IPN nanogel), in which lipase is encapsulated into an interpenetrating polymer matrix formed by polyacrylamide and poly(N-isopropylacrylamide) (PNIPAAm) has been designed and synthesized for an enhanced stability and activity in both aqueous and non-polar organic solvents. A three-step method, including acryloylation, polymerization with acrylamide and sequential polymerization with N-isopropylacrylamide, was established to fabricate enzyme nanogel with temperature-sensitive interpenetrating polymer network. It has been shown by an all-atom molecular dynamics simulation that above mentioned polymer matrix forms a more hydrophobic environment, as compared to that obtained with sole polyacrylamide, because of the penetration of N-isopropylacrylamide into the polymer acrylamide network via hydrogen bonding, which is further confirmed by the fluorescence spectrum. This favours the uptake of hydrophobic substrates and thus the overall rate of enzymatic catalysis. The enhanced stability and catalytic performance of this novel lipase nanogel in aqueous and non-polar organic solvent were demonstrated by using hydrolysis reaction of p-NPP in aqueous and esterification reaction of ibuprofen in isooctane. In aqueous solution, the residual activity of CRL-IPN nanogel maintains its 70% activity at 60 °C after 4 h, compared with that free lipase only has 30% at the same condition. In addition, the CRL-IPN nanogel can be reused for 10 cycles with no loss of its activity. In isooctane, CRL-IPN nanogel gave a 33% yield of esterification of ibuprofen, in comparison to 22% using free lipase and less than 5% using lipase encapsulated in a polyacrylamide matrix. The enhanced stability and activity make this CRL-IPN nanogel promising for enzymatic catalysis in non-polar solvents.Graphical abstractDownload full-size imageHighlights► We fabricated lipase nanogel with interpenetrating polymer network. ► This lipase nanogel can be recycled due to its temperature-responsive property. ► This lipase nanogel can catalyze reaction in non-polar organic solvent. ► The reason of forming interpenetrating polymer network is provided by using molecular dynamic simulation. ► The mechanism of catalysis of reaction in non-polar organic solvent is proposed by using molecular dynamic simulation.
Co-reporter:Kai KANG, Diannan LU, Zheng LIU
Chinese Journal of Chemical Engineering (April 2012) Volume 20(Issue 2) pp:284-293
Publication Date(Web):1 April 2012
DOI:10.1016/S1004-9541(12)60390-5
Poly(N-isopropylacrylamide) (PNIPAAm) grafted onto silica, which may be used for reverse phase chromatography (RPC), was simulated and synthesized for protein separation with temperature-triggered adsorption and desorption. Molecular dynamics simulation at an all-atom level was performed to illustrate the adsorption/desorption behavior of cytochrome c, the model protein, on PNIPAAm-grafted-silica, a temperature responsive adsorbent. At a temperature above the lower critical solution temperature (LCST), the PNIPAAm chains aggregate on the silica surface, forming a hydrophobic surface that is favorable for the hydrophobic adsorption of cytochrome c, which has a high exposure of hydrophobic patches. At temperatures below the LCST, the PNIPAAm chains stretch, forming hydrophilic surface due to hydrogen bonding between PNIPAAm and surrounding water. Desorption of cytochrome c on the PNIPAAm-grafted-silica surface occurs as a result of competition with water, which forms hydrogen bonds with the protein. The conformational transitions of both cytochrome c and PNIPAAm are monitored, providing molecular insight into this temperature-responsive RPC technique. PNIPAAm-grafted-silica beads were synthesized and used for the adsorption and desorption of cytochrome c at approximately 313 K and 290 K, respectively. The experimental results validate the molecular dynamics simulation. In comparison to conventional RPC, using temperature as a driving force for RPC reduces the risk of protein denaturation caused by exposure to chaotropic solvents. Moreover, it simplifies the separation process by avoiding the buffer exchange operations between the steps.
Co-reporter:Xiufu Hua, Jun Wang, Zuojun Wu, Hongxing Zhang, Heping Li, Xinghui Xing, Zheng Liu
Biochemical Engineering Journal (15 April 2010) Volume 49(Issue 2) pp:201-206
Publication Date(Web):15 April 2010
DOI:10.1016/j.bej.2009.12.014
Co-reporter:Jun Ge, Diannan Lu, Zhixia Liu, Zheng Liu
Biochemical Engineering Journal (15 April 2009) Volume 44(Issue 1) pp:53-59
Publication Date(Web):15 April 2009
DOI:10.1016/j.bej.2009.01.002
Co-reporter:Kun Zhang, Yuanyuan Xu, Xiufu Hua, Huilong Han, Jiannan Wang, Jun Wang, Yongmin Liu, Zheng Liu
Biochemical Engineering Journal (1 October 2008) Volume 41(Issue 3) pp:251-257
Publication Date(Web):1 October 2008
DOI:10.1016/j.bej.2008.05.003
Co-reporter:Gong Chen, Xian Kong, Diannan Lu, Jianzhong Wu and Zheng Liu
Physical Chemistry Chemical Physics 2017 - vol. 19(Issue 18) pp:NaN11697-11697
Publication Date(Web):2017/04/07
DOI:10.1039/C7CP00887B
Molecular dynamics (MD) simulations, in combination with the Markov-state model (MSM), were applied to probe CO2 diffusion from an aqueous solution into the active site of human carbonic anhydrase II (hCA-II), an enzyme useful for enhanced CO2 capture and utilization. The diffusion process in the hydrophobic pocket of hCA-II was illustrated in terms of a two-dimensional free-energy landscape. We found that CO2 diffusion in hCA-II is a rate-limiting step in the CO2 diffusion-binding-reaction process. The equilibrium distribution of CO2 shows its preferential accumulation within a hydrophobic domain in the protein core region. An analysis of the committors and reactive fluxes indicates that the main pathway for CO2 diffusion into the active site of hCA-II is through a binding pocket where residue Gln136 contributes to the maximal flux. The simulation results offer a new perspective on the CO2 hydration kinetics and useful insights toward the development of novel biochemical processes for more efficient CO2 sequestration and utilization.
Co-reporter:Yifei Zhang, Qile Chen, Jun Ge and Zheng Liu
Chemical Communications 2013 - vol. 49(Issue 84) pp:NaN9817-9817
Publication Date(Web):2013/08/29
DOI:10.1039/C3CC45837G
An enzyme-incorporated hydrogel made of alginate and polyacrylamide shows a linearly increased activity with the enlargement of surface area upon stretching.
Co-reporter:Xian Kong, Shanshan Qin, Diannan Lu and Zheng Liu
Physical Chemistry Chemical Physics 2014 - vol. 16(Issue 18) pp:
Publication Date(Web):
DOI:10.1039/C3CP55524K
Co-reporter:Binbin Wu, Tian Lan, Diannan Lu and Zheng Liu
Environmental Science: Nano 2014 - vol. 16(Issue 6) pp:NaN1509-1509
Publication Date(Web):2014/02/28
DOI:10.1039/C3EM00731F
The changes in microbial ecology interpreted from taxonomic and functional genes and biological functions represented by urease and dehydrogenase activities were monitored in soil contaminated with different petroleum hydrocarbons including crude oil, diesel, n-hexadecane and poly-aromatic hydrocarbons (PAHs). It was shown that the presence of n-hexadecane stimulated the activity of indigenous microorganisms, especially alkane degrading bacteria, and led to over 20% degradation of n-hexadecane within one month. No obvious degradation of the other three types of petroleum hydrocarbons was observed. The stimulation effect was most marked in the soil spiked with a medium concentration (2500 mg kg−1 dry soil) of n-hexadecane. However, the presence of PAHs completely inhibited the previously-mentioned bioactivities of the soil. The content of PAH degrading bacteria, however, increased more than 10-fold, indicating the selection effect of PAHs on soil bacteria. The impacts of diesel and crude oil on the microbial ecology and biological functions varied significantly with their concentration. The disclosure of the ecological and enzymatic responses could be helpful in soil bioremediation.
Co-reporter:Jingying Zhu, Yifei Zhang, Diannan Lu, Richard N. Zare, Jun Ge and Zheng Liu
Chemical Communications 2013 - vol. 49(Issue 54) pp:NaN6092-6092
Publication Date(Web):2013/05/22
DOI:10.1039/C3CC42493F
A general approach for preparing enzyme–polymer nanoconjugates that respond to temperature in organic media is presented. These nanoconjugates readily dissolve in organic solvents for homogenous catalysis at 40 °C and showed greatly enhanced apparent catalytic activities. The recovery of the soluble enzyme–polymer nanoconjugates is accomplished by temperature-induced precipitation.
Co-reporter:Yifei Zhang, Fengjiao Lyu, Jun Ge and Zheng Liu
Chemical Communications 2014 - vol. 50(Issue 85) pp:NaN12922-12922
Publication Date(Web):2014/08/28
DOI:10.1039/C4CC06158F
A method using ink-jet printing for constructing multi-enzyme systems was proposed, in which a precisely defined enzyme ratio and two-dimensional distribution was obtained by the preset ‘color’ values. The applications of the print-on-paper multi-enzyme systems were exemplified by the detection of glucose and the design of an enzyme-enabled two-dimensional code.
Co-reporter:Zhixia Liu, Diannan Lu, Jianmin Li, Wei Chen and Zheng Liu
Physical Chemistry Chemical Physics 2009 - vol. 11(Issue 2) pp:NaN340-340
Publication Date(Web):2008/11/24
DOI:10.1039/B811496J
The aim of this study was to obtain molecular insight into the deactivation of recombinant urate oxidase (uricase, UOX, EC 1.7.3.3) (rUOX) from Aspergillus flavus. The enzyme is a tunnel-shaped homotetramer and has important clinical applications. By means of molecular dynamics simulations, multidimensional structural characterization and enzyme activity assays, we concluded that the thermal deactivation of UOX at neutral pH was associated with the loss of intersubunit hydrogen (H) bonds. This mechanism could also explain the deactivation of dilute aqueous UOX. Thermal deactivation of aqueous UOX due to dissociation of its subunits was ruled out. Displacement of H2O from the surface of UOX by less polar solvents such as methanol and dimethyl sulfoxide (DMSO) was proposed as an approach for strengthening intersubunit H bonds and consequently UOX stability. The effectiveness of this method was validated by both in silico and in vitro experiments. The results mentioned above provide insights for improving the stability of UOX and extending its applications. They may also be helpful for understanding the properties of other multimeric proteins.
Co-reporter:Yijie Dong, Zhe Lang, Xian Kong, Diannan Lu and Zheng Liu
Environmental Science: Nano 2015 - vol. 17(Issue 4) pp:NaN774-774
Publication Date(Web):2015/02/10
DOI:10.1039/C4EM00428K
Biostimulation, which employs nutrients to enhance the proliferation of indigenous microorganisms and therefore the degradation of contaminants, is an effective tool for treatment of oil-contaminated soil. However, the evolution of microbial ecology, which responds directly to stimulation procedures and intrinsically determines the degradation of oil contaminants, has rarely been explored, particularly in the context of biostimulation. In this study, the effects of biostimulation procedures including the regulation of the C:N:P ratio, as well as application of surfactants and electron acceptors in the degradation of crude oil contaminants and the evolution of the microbial community were examined simultaneously to provide ecological insights into the biostimulation. The real-time PCR showed that biostimulation promoted the proliferation of bacteria, with Gammaproteobacteria showing the greatest increase. However, the proliferation of fungi was inhibited by the accumulation of the degradation products. The degradation of polar compounds of crude oil contaminants was characterized by negative-ion electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (negative-ion ESI FT-ICR MS), showing a biased increase in the relative abundance of naphthenic acids. Principal component analysis (PCA) showed that different species in oil sludge have different degradation rates during biostimulation. The addition of fertilizers with surfactants and electron acceptors profoundly stimulated the indigenous microorganisms with N1, O1 and O2 species as substrates while those with O3 and O4 species were little affected. An enriched abundance of alkB genes was observed during the degradation of saturated hydrocarbons. Monitoring the kinetics of the microbial community, functional genes and degradation offers a comprehensive view for the understanding and optimization of the biostimulation process.
Co-reporter:Gong Chen, Xian Kong, Jingying Zhu, Diannan Lu and Zheng Liu
Physical Chemistry Chemical Physics 2015 - vol. 17(Issue 16) pp:NaN10714-10714
Publication Date(Web):2015/03/12
DOI:10.1039/C5CP00418G
While the conjugation of enzymes with ABA copolymers has resulted in increased enzymatic activities in organic solvents, by several orders of magnitude, the underpinning mechanism has not been fully uncovered, particularly at the molecular level. In the present work, a coarse-grained molecular dynamics simulation of cytochrome c (Cyt c) conjugated with a PEO–PPO–PEO block copolymer (ABA) in toluene was simulated with Cyt c as a control. It is shown that the hydrophilic segments (PEO) of the conjugated block copolymer molecules tend to entangle around the hydrophilic patch of Cyt c, while the hydrophobic segments (PPO) extend into the toluene. At a lower temperature, the PEO tails tend to form a hairpin structure outside the conjugated protein, whereas the Cyt c–ABA conjugates tend to form larger aggregates. At a higher temperature, however, the PEO tails tend to adsorb onto the hydrophilic protein surface, thus improving the suspension of the Cyt c–ABA conjugates and, consequently, the contact with the substrate. Moreover, the temperature increase drives the conformational transition of the active site of Cyt c–ABA from an “inactive state” to an “activated state” and thus results in an enhanced activity. To validate the above simulations, Cyt c was conjugated to F127, an extensively used ABA copolymer. By elevating the temperature, a decrease in the average size of the Cyt c–F127 conjugates along with a great increase in the apparent activity in toluene was observed, as can be predicted from the molecular dynamics simulation. The above mentioned molecular simulations offer a molecular insight into the temperature-responsive behaviour of protein–ABA copolymers, which is helpful for the design and application of enzyme–polymer conjugates for industrial biocatalysis.
Co-reporter:Zuojun Wu, Liangdong Zou, Diannan Lu and Zheng Liu
Environmental Science: Nano 2011 - vol. 13(Issue 10) pp:NaN2913-2913
Publication Date(Web):2011/09/08
DOI:10.1039/C0EM00761G
Soil microbial ecosystems are responsive to environmental changes that underpin the biological functions of the soil. The present study was conducted to profile variations in the microbial ecological system of remediated soil (R) and petroleum contaminated soil (P) based on comparisons with soil that had not been contaminated (N), using a cloning library of taxonomic genes (16S rRNA gene for bacteria and 18S rRNA gene for eukaryotes) and functional genes (nifH, amoA and narG). The results showed that N and R had a similar distribution in both the taxonomic genes and functional genes for bacteria and eukaryotes, which were dominated by Proteobacteria and Arthropoda, respectively. Phylogenetic analysis based on the nifHgene showed that the sequences from the three soils were clustered into six taxonomic groups, Actinobacteridae, and Alpha-, Beta-, Gamma- and Delta-proteobacteria, as well as an unclassified group. Evaluation of the amoAgene revealed that all sequences derived from the three samples belonged to Betaproteobacteria. The R and N soil had similar Shannon–Wiener diversity index (H′) values, both of which were significantly higher than that of the P soil. The most abundant bacterial phylotype identified in the N and R soils were the same and were related to an uncultured bacterial clone (GAN-SB17, FN423475). None of the narGgenes were found in the P soil. Similar results in terms of distribution, composition and the related index were obtained for nifH and amoA. These parameters may comprise a biological ecology index that may be applied to aid the design, implementation and evaluation of soil bioremediation.
Co-reporter:Mengmeng Lin, Diannan Lu, Jingying Zhu, Cheng Yang, Yifei Zhang and Zheng Liu
Chemical Communications 2012 - vol. 48(Issue 27) pp:NaN3317-3317
Publication Date(Web):2012/02/13
DOI:10.1039/C2CC30189J
A universal synthetic route for magnetic enzyme nanogels (MENGs) was proposed, based on electrostatic interaction driven assembly and in situ polymerization from the surface of magnetic nanoparticles, to avoid chemical modification of proteins and hence structural and functional deterioration.
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