Temperature-triggered Protein Adsorption and Desorption on Temperature-responsive PNIPAAm-grafted-silica:Molecular Dynamics Simulation and Experimental Validation

Abstract

Abstract: 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.

Key words: reverse phase chromatography, PNIPAAm-grafted-silica, cytochrome c, molecular dynamics simulation, atom transfer radical polymerization (ATRP)

摘要： 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.

关键词: reverse phase chromatography, PNIPAAm-grafted-silica, cytochrome c, molecular dynamics simulation, atom transfer radical polymerization (ATRP)

KANG Kai, LU Diannan, LIU Zheng. Temperature-triggered Protein Adsorption and Desorption on Temperature-responsive PNIPAAm-grafted-silica:Molecular Dynamics Simulation and Experimental Validation[J]. Chin.J.Chem.Eng., 2012, 20(2): 284-293.

康锴, 卢滇楠, 刘铮. Temperature-triggered Protein Adsorption and Desorption on Temperature-responsive PNIPAAm-grafted-silica:Molecular Dynamics Simulation and Experimental Validation[J]. Chinese Journal of Chemical Engineering, 2012, 20(2): 284-293.

References

1 Janson,J.C.,“The development of gel media and columns for large-scale chromatography of proteins,a historical review”,Chin.J. Chem.Eng.,10(6),690-695(2002).
2 Ivanov,A.E.,Galaev,I.Y.,Kazakov,S.V.,Mattiasson,B.,“Thermosensitive copolymers of N-vinylimidazole as displacers of proteins in immobilized metal affinity chromatography”,J.Chromatogr.A,907 (1/2),115-130(2001).
3 Schild,H.G.,“Poly(N-isopropylacrylamide):experiment,theory and application”,Progress in Polymer Science,17(2),163-249(1992).
4 Lu,D.N.,Liu,Z.X.,Zhang,M.L.,Wang,X.G.,Liu,Z.,“Dextrangrafted-PNIPAAm as an artificial chaperone for protein refolding”, Biochem.Eng.J.,27,336-343(2006).
5 Kanazawa,H.,Nishikawa,M.,Mizutani,A.,Sakamoto,C.,MoritaMurase,Y.,Nagata,Y.,Kikuchi,A.,Okano,T.,“Aqueous chromatographic system for separation of biomolecules using thermoresponsive polymer modified stationary phase”,J.Chromatogr.A,1191(2),157-161(2008).
6 Maharjan,P.,Hearn,M.T.W.,Jackson,W.R.,“Development of a temperature-responsive agarose-based ion-exchange chromatographic resin”,J.Chromatogr.A,1216(50),8722-8729(2009).
7 Akiyama,Y.,Shinohara,Y.,Hasegawa,Y.,Kikuchi,A.,Okano,T., “Preparation of novel acrylamide-based thermoresponsive polymer analogues and their application as thermoresponsive chromatographic matrices”,J.Polym.Sci.A—Polym.Chem.,46(16),5471-5482(2008).
8 Kidoaki,S.,Ohya,S.,Nakayama,Y.,Matsuda,T.,“Thermoresponsive structural change of a poly(N-isopropylacrylamide)graft layer measured with an atomic force microscope”,Langmuir,17(8),2402-2407(2001).
9 Xu,F.J.,Zhong,S.P.,Yung,L.Y.L.,Kang,E.T.,Neoh,K.G.,“Surface-active and stimuli-responsive polymer-Si(100)hybrids from surface-initiated atom transfer radical polymerization for control of cell adhesion”,Biomacromolecules,5(6),2392-2403(2004).
10 Fu,Q.,Rao,G.V.R.,Basame,S.B.,Keller,D.J.,Artyushkova,K., Fulghum,J.E.,Lopez,G.P.J.,“Reversible control of free energy and topography of nanostructured surfaces”,Journal of the American Chemical Society,126(29),8904-8905(2004).
11 Balamurugan,S.,Mendez,S.,Balamurugan,S.S.,O’Brien,M.J., Lopez,G.P.,“Thermal response of poly(N-isopropylacrylamide) brushes probed by surface plasmon resonance”,Langmuir,19(7),2545-2549(2003).
12 Takei,Y.G.,Aoki,T.,Sanui,K.,Ogata,N.,Sakurai,Y.,Okano,T., “Dynamic contact-angle measurement of temperature-responsive surface properties for poly(N-isopropylacrylamide)grafted surfaces”, Macromolecules,27(21),6163-6166(1994).
13 Jones,D.M.,Smith,J.R.,Huck,W.T.S.,Alexander,C.,“Variable adhesion of micropatterned thermoresponsive polymer brushes: AFM investigations of poly(N-isopropylacrylamide)brushes prepared by surface-initiated polymerizations”,Advanced Materials,14 (16),1130-1134(2002).
14 Lindqvist,J.,Nystrom,D.,Ostmark,E.,“Intelligent dual-responsive cellulose surfaces via surface-initiated ATRP”,Biomacromolecules,9,2139-2145(2008).
15 Lin,S.Y.,Chen,K.S.,Run-Chu,L.,“Thermal micro ATR/FT-IR spectroscopic system for quantitative study of the molecular structure of poly(N-isopropylacrylamide)in water”,Polymer,40(10),2619-2624(1999).
16 Longhi,G.,Lebon,F.,Abbate,S.,“Molecular dynamics simulation of a model oligomer for poly(N-isopropylamide)in water”,Chemical Physics Letters,386,123-127(2004).
17 Gangemi,F.,Longhi,G.,Abbate,S.,Lebon,F.,Cordone,R., Ghilardi,G.P.,Fornili,S.L.,“Molecular dynamics simulation of aqueous solutions of 26-unit segments of p(NIPAAm)and of p(NIPAAm)‘Doped’with amino acid based comonomers”,J.Phys. Chem.B,112(38),11896-11906(2008).
18 Lu,D.N.,Liu,Z.,“Molecular simulation of polymer assisted protein refolding”,J.Chem.Phy.,123(13),134903(2005).
19 Giovambattista,N.,Rossky,P.J.,Debenedetti,P.G.,“Effect of pressure on the phase behavior and structure of water confined between nanoscale hydrophobic and hydrophilic plates”,Physical Review E,73(4),041604(2006).
20 Garcia-Arellano,H.,Valderrama,B.,Saab-Rincon,G.,VazquezDuhalt,R.,“High temperature biocatalysis by chemically modified cytochrome c”,Bioconjugate Chemistry,13(6),1336-1344(2002).
21 Berendsen,H.J.C.,Grigera,J.R.,Stroatsma,T.P.,“The missing term in effective pair potentials”,J.Phys.Chem.,91,6269-6271(1987).
22 Spoel,D.,Lindahl,E.,Hess,B.,Buuren,A.R.,Apol,E.,Meulenhoff, P.J.,Tieleman,D.P.,Sijbers,A.L.T.M.,Feenstra,K.A.,Drunen,R., Berendsen,H.J.C.,Gromacs User Manual version 4.5,www.gromacs.org (2010).
23 Galaev,I.Y.,Mattiasson,B.,“Smart polymers and what they could do in biotechnology and medicine”,Trends in Biotechnology,17(8),335-340(1999).
24 Wang,X.,Xiao,X.,Wang,X.,Zhou,J.,Li,L.,Xu,J.,Guo,B., “Reversibly switchable double-responsive block copolymer brushes”, Macromolecular Rapid Communications,28(7),828-833(2007).
25 Kang,K.,Lu,D.N.,Zhang,M.L.,Liu,Z.,“All-atom molecular dynamics simulation of protein separation process by reverse phase liquid chromatography”,CISEC J.,3,660-667(2010).(in Chinese)

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