分子动力学模拟研究穿膜肽bLFcin6与不同磷脂双层膜的相互作用
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摘要
穿膜肽是一类由5~30个氨基酸残基组成的有穿膜能力的小分子多肽,可携带各种物质进入细胞内部,发挥各自生物学活性.bLFcin6是一种新的细胞穿透肽,选取含有4~9个残基的bLFcin6为研究对象,用Pymol软件来构建肽的初始结构并下载膜DPPC,POPC,POPG的结构,在优化后搭建肽-膜的初始体系.每个体系模拟了500ns,对肽用GROMOS53a6力场处理,对膜用Berger力场处理,温度设置为323K.在这3个体系中,bLFcin6进入到DPPC的羰基端并与疏水的尾部相互作用;bLFcin6能进入到POPG的亲水头部,不能更深的插入到内部;bLFcin6并不能接触到膜上.分析了在不同氨基酸与膜带电基团之间氢键的形成,发现了带正电的精氨酸在肽-膜之间的相互作用起了一个重要的角色.计算了在不同的氨基酸和膜的质心之间的距离,发现肽的N端首先接触到DPPC和POPG的膜上.此外,不同膜对bLFcin6穿膜能力的影响与脂质分子的结构有关.BLFcin6对不同膜有不同的渗透能力.我们重点强调了精氨酸的重要性,与先前实验和模拟中得到的结果是一致的,这为进一步探究bLFcin6的穿膜机制提供了一定的帮助.
Cell Penetrating Peptides are a kind of small molecular peptides.These peptides are composed of 5~30amino acid residues with the membrane penetration ability and can carry a variety of material into the cell.In this study,we selected lactoferricin B,4~9residue RRWQWR(bLFcin6),which is a newly discovered cell penetrating peptide.We used Pymol software to build the initial structure of peptide and downloaded DPPC,POPC and POPG membrane from website,after the optimization,built peptidemembrane systems.Three 500 ns long independent molecular dynamics simulations were performed with Gromacs 4.5.3software.The GROMOS53a6 force field was adopted for peptide and Berger force field for membrane.The temperature was set at 323 K.For the three kinds of membrane simulation,bLFcin6 entered into DPPC membrane to the carbonyl group and interacted with the hydrophobic tail;bLFcin6interacted with the hydrophilic head group of POPG membrane,not into the deeper;bLFcin6did not contacted with POPC membrane.We analyzed the hydrogen bonds formation between different amino acids and membrane phosphate group,found that positively charged arginine plays an important role in peptidemembranes interaction.Also,we calculated the distance between different amino acids and the center-of-mass(COM)of membrane,obtained that the N end of peptide first contacted with the DPPC and POPG membranes.In addition,the influence of different membranes to bLFcin6 is associated with the structure of lipid molecules.BLFcin6 have the different penetration ability to different bilayer membranes.Here,it the importance of arginine is highlighted in order to consistent with previous biological experiment and simulation analysis,it provides the reference for the further analysis of the penetration membrane process.
Cell Penetrating Peptides are a kind of small molecular peptides.These peptides are composed of 5~30amino acid residues with the membrane penetration ability and can carry a variety of material into the cell.In this study,we selected lactoferricin B,4~9residue RRWQWR(bLFcin6),which is a newly discovered cell penetrating peptide.We used Pymol software to build the initial structure of peptide and downloaded DPPC,POPC and POPG membrane from website,after the optimization,built peptidemembrane systems.Three 500 ns long independent molecular dynamics simulations were performed with Gromacs 4.5.3software.The GROMOS53a6 force field was adopted for peptide and Berger force field for membrane.The temperature was set at 323 K.For the three kinds of membrane simulation,bLFcin6 entered into DPPC membrane to the carbonyl group and interacted with the hydrophobic tail;bLFcin6interacted with the hydrophilic head group of POPG membrane,not into the deeper;bLFcin6did not contacted with POPC membrane.We analyzed the hydrogen bonds formation between different amino acids and membrane phosphate group,found that positively charged arginine plays an important role in peptidemembranes interaction.Also,we calculated the distance between different amino acids and the center-of-mass(COM)of membrane,obtained that the N end of peptide first contacted with the DPPC and POPG membranes.In addition,the influence of different membranes to bLFcin6 is associated with the structure of lipid molecules.BLFcin6 have the different penetration ability to different bilayer membranes.Here,it the importance of arginine is highlighted in order to consistent with previous biological experiment and simulation analysis,it provides the reference for the further analysis of the penetration membrane process.
引文
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[13]van Buuren,A.R.,Marrink,S.J.,Berendsen,H.J..A molecular dynamics study of the decane/water interface[J].The Journal of Physical Chemistry,1993,97(36):9206-9212.
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[18]Herce,H.D.,Garcia,A.E..Molecular dynamics simulations suggest a mechanism for translocation of the HIV-1TAT peptide across lipid membranes[J].Proceedings of the National Academy of Sciences of the United States of America,2007,104(52):20805-10.
[19]Van Der Spoel,D.,Lindahl,E.,Hess,B.,Groenhof,G.,Mark,A.E.,Berendsen,H.J..GROMACS:fast,flexible,and free[J].J Comput Chem,2005,26(16):1701-18.
[20]Herce,H.,Garcia,A.,Litt,J.,Kane,R.,Martin,P.,Enrique,N.,Rebolledo,A.,Milesi,V..Arginine-rich peptides destabilize the plasma membrane,consistent with a pore formation translocation mechanism of cell-penetrating peptides[J].Biophysical journal,2009,97(7):1917-1925.
[21]Hess,B.,Kutzner,C.,Van Der Spoel,D.,Lindahl,E..GROMACS 4:algorithms for highly efficient,load-balanced,and scalable molecular simulation[J].Journal of chemical theory and computation,2008,4(3):435-447.
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[23]Berger,O.,Edholm,O.,Jhnig,F..Molecular dynamics simulations of a fluid bilayer of dipalmitoylphosphatidylcholine at full hydration,constant pressure,and constant temperature[J].Biophysical journal,1997,72(5):2002.
[24]Berendsen,H.J.;Postma,J.P.;van Gunsteren,W.F.;Hermans,J..Interaction models for water in relation to protein hydration[J].In Intermolecular forces,Springer,1981:331-342.
[25]Schrodinger,L..The PyMOL molecular graphics system,version 1.3r1.2010.There is no corresponding record for this reference 2010.
[26]Saini,A.,R.Jaswal,R.,Negi,R.,S.Nandel,F..Insights on the structural characteristics of Vim-TBS(58-81)peptide for future applications as a cell penetrating peptide[J].Bioscience trends,2013,7(5):209-220.
[27]Pourmousa,M.,Wong-ekkabut,J.,Patra,M.,Karttunen,M..Molecular dynamic studies of transportan interacting with a DPPC lipid bilayer[J].The journal of physical chemistry,B 2013,117(1):230-41.
[28]Song,J.,Kai,M.,Zhang,W.,Zhang,J.,Liu,L.,Zhang,B.,Liu,X.,Wang,R..Cellular uptake of transportan 10and its analogs in live cells:Selectivity and structure-activity relationship studies[J].Peptides,2011,32(9):1934-41.
[29]Dunkin,C.M.,Pokorny,A.,Almeida,P.F.,Lee,H.S..Molecular dynamics studies of transportan 10(tp10)interacting with a POPC lipid bilayer[J].The journal of physical chemistry.B,2011,115(5):1188-98.
[30]Kawamoto,S.,Takasu,M.,Miyakawa,T.,Morikawa,R.,Oda,T.,Futaki,S.,Nagao,H..Binding of Tat peptides on DOPC and DOPG lipid bilayer membrane studied by molecular dynamics simulations[J].Molecular Simulation,2012,38(5):366-368.
[31]Kim,K.,Jordan,K..Particle mesh Ewald:An N[center-dot]log(N)method for Ewald sums in large systems[J].J.Chem.Phys,1994,98:10089.
[32]Hess,B.,Bekker,H.,Berendsen,H.J.,Fraaije,J.G..LINCS:a linear constraint solver for molecular simulations[J].Journal of computational chemistry,1997,18(12):1463-1472.
[33]张继伟,卞富永,施国军,徐四川.多巴胺在POPC磷脂双层膜中扩散和透过过程的分子动力学模拟[J].Acta Phys.-Chim.Sin,2014:30.
[34]张贇.人胰岛淀粉样多肽hIAPP单体及二聚体与POPG磷脂膜相互作用的分子动力学模拟研究[D].上海:复旦大学,2012.
[35]常钟文.淀粉样Aβ25-35片段寡聚体与POPG双层膜相互作用的分子动力学模拟[D].上海:复旦大学,2010.
[36]Fimland,G.,Eijsink,V.G.,Nissen-Meyer,J..Mutational analysis of the role of tryptophan residues in an antimicrobial peptide[J].Biochemistry,2002,41(30):9508-9515.
[37]Derossi,D.,Chassaing,G.,Prochiantz,A..Trojan peptides:the penetratin system for intracellular delivery[J].Trends in cell biology,1998,8(2):84-87.
[38]Richard,J.P.,Melikov,K.,Vives,E.,Ramos,C.,Verbeure,B.,Gait,M.J..Chernomordik,L.V.,Lebleu,B..Cell-penetrating peptides A reevaluation of the mechanism of cellular uptake[J].Journal of Biological Chemistry,2003,278(1):585-590.
[2]陈翀,季守平.穿膜肽的研究现状及应用[J].生物技术通讯,2012(1):142-147.
[3]曹赞霞,刘磊,王吉华.分子动力学模拟研究穿透肽的跨膜机制及引导新肽设计[J].科学通报(中文版)2014,59(22),2160-2168.
[4]Hilchie,A.L.,Vale,R.,Zemlak,T.S.,Hoskin,D.W..Generation of a hematologic malignancy-selective membranolytic peptide from the antimicrobial core(RRWQWR)of bovine lactoferricin[J].Experimental and molecular pathology,2013,95(2):192-8.
[5]Ammons,M.C.,Copie,V..Mini-review:Lactoferrin:a bioinspired,anti-biofilm therapeutic[J].Biofouling,2013,29(4):443-55.
[6]Duchardt,F.,Ruttekolk,I.R.,Verdurmen,W.P.,Lortat-Jacob,H.,Burck,J.,Hufnagel,H.,Fischer,R.,van den Heuvel,M.,Lowik,D.W.,Vuister,G.W.,Ulrich,A.,de Waard,M.,Brock,R..A cell-penetrating peptide derived from human lactoferrin with conformation-dependent uptake efficiency[J].J Biol Chem2009,284(52):36099-108.
[7]Fang,B.,Guo,H.Y.,Zhang,M.,Jiang,L.,Ren,F.Z..The six amino acid antimicrobial peptide bLFcin6penetrates cells and delivers siRNA[J].The FEBS journal,2013,280(4):1007-17.
[8]Witte,K.,Olausson,B.E.,Walrant,A.,Alves,I.D.,Vogel,A..Structure and dynamics of the two amphipathic arginine-rich peptides RW9and RL9in a lipid environment investigated by solid-state NMR and MD simulations[J].Biochimica et biophysica acta,2013,1828(2):824-33.
[9]Walrant,A.,Vogel,A.,Correia,I.,Lequin,O.,Olausson,B.E S.,Desbat,B.,Sagan,S.,Alves,I.D..Membrane interactions of two arginine-rich peptides with different cell internalization capacities[J].Biochimica et Biophysica Acta(BBA)-Biomembranes,2012,1818(7):1755-1763.
[10]MacKerell,A.D.,Feig,M.,Brooks,C.L..Improved treatment of the protein backbone in empirical force fields[J].Journal of the American Chemical Society,2004,126(3):698-699.
[11]MacKerell,A.D.,Bashford,D.,Bellott,M.,Dunbrack,R.,Evanseck,J.,Field,M.J.,Fischer,S.,Gao,J.,Guo,H.,Ha,S.a..All-atom empirical potential for molecular modeling and dynamics studies of proteins[J].The journal of physical chemistry B,1998,102(18):3586-3616.
[12]Pourmousa,M.,Karttunen,M..Early stages of interactions of cell-penetrating peptide penetratin with a DPPC bilayer[J].Chemistry and physics of lipids,2013,169:85-94.
[13]van Buuren,A.R.,Marrink,S.J.,Berendsen,H.J..A molecular dynamics study of the decane/water interface[J].The Journal of Physical Chemistry,1993,97(36):9206-9212.
[14]Mark,A.E.,van Helden,S.P.,Smith,P.E.,Janssen,L.H.,van Gunsteren,W.F..Convergence Properties Of Free Energy Calculations:alpha.-cyclodextrin complexes as a case study[J].Journal of the American Chemical Society,1994,116(14):6293-6302.
[15]Lensink,M.,Christiaens,B.,Vandekerckhove,J.,Prochiantz,A.;Rosseneu,M..Penetratin-membrane association:W48/R52/W56shield the peptide from the aqueous phase[J].Biophysical journal,2005,88(2):939-952.
[16]van Gunsteren,W.F.,Billeter,S.,Eising,A.,Hünenberger,P.H.,Krüger,P.,Mark,A.E.,Scott,W.,Tironi,I.G..Biomolecular simulation:The{GROMOS96}manual and user guide,1996.
[17]Yesylevskyy,S.,Marrink,S.J.,Mark,A.E..Alternative mechanisms for the interaction of the cell-penetrating peptides penetratin and the TAT peptide with lipid bilayers[J].Biophysical journal,2009,97(1):40-9.
[18]Herce,H.D.,Garcia,A.E..Molecular dynamics simulations suggest a mechanism for translocation of the HIV-1TAT peptide across lipid membranes[J].Proceedings of the National Academy of Sciences of the United States of America,2007,104(52):20805-10.
[19]Van Der Spoel,D.,Lindahl,E.,Hess,B.,Groenhof,G.,Mark,A.E.,Berendsen,H.J..GROMACS:fast,flexible,and free[J].J Comput Chem,2005,26(16):1701-18.
[20]Herce,H.,Garcia,A.,Litt,J.,Kane,R.,Martin,P.,Enrique,N.,Rebolledo,A.,Milesi,V..Arginine-rich peptides destabilize the plasma membrane,consistent with a pore formation translocation mechanism of cell-penetrating peptides[J].Biophysical journal,2009,97(7):1917-1925.
[21]Hess,B.,Kutzner,C.,Van Der Spoel,D.,Lindahl,E..GROMACS 4:algorithms for highly efficient,load-balanced,and scalable molecular simulation[J].Journal of chemical theory and computation,2008,4(3):435-447.
[22]Oostenbrink,C.,Soares,T.A.,Van der Vegt,N.F.,Van Gunsteren,W.F..Validation of the 53A6 GROMOS force field[J].European Biophysics Journal,2005,34(4):273-284.
[23]Berger,O.,Edholm,O.,Jhnig,F..Molecular dynamics simulations of a fluid bilayer of dipalmitoylphosphatidylcholine at full hydration,constant pressure,and constant temperature[J].Biophysical journal,1997,72(5):2002.
[24]Berendsen,H.J.;Postma,J.P.;van Gunsteren,W.F.;Hermans,J..Interaction models for water in relation to protein hydration[J].In Intermolecular forces,Springer,1981:331-342.
[25]Schrodinger,L..The PyMOL molecular graphics system,version 1.3r1.2010.There is no corresponding record for this reference 2010.
[26]Saini,A.,R.Jaswal,R.,Negi,R.,S.Nandel,F..Insights on the structural characteristics of Vim-TBS(58-81)peptide for future applications as a cell penetrating peptide[J].Bioscience trends,2013,7(5):209-220.
[27]Pourmousa,M.,Wong-ekkabut,J.,Patra,M.,Karttunen,M..Molecular dynamic studies of transportan interacting with a DPPC lipid bilayer[J].The journal of physical chemistry,B 2013,117(1):230-41.
[28]Song,J.,Kai,M.,Zhang,W.,Zhang,J.,Liu,L.,Zhang,B.,Liu,X.,Wang,R..Cellular uptake of transportan 10and its analogs in live cells:Selectivity and structure-activity relationship studies[J].Peptides,2011,32(9):1934-41.
[29]Dunkin,C.M.,Pokorny,A.,Almeida,P.F.,Lee,H.S..Molecular dynamics studies of transportan 10(tp10)interacting with a POPC lipid bilayer[J].The journal of physical chemistry.B,2011,115(5):1188-98.
[30]Kawamoto,S.,Takasu,M.,Miyakawa,T.,Morikawa,R.,Oda,T.,Futaki,S.,Nagao,H..Binding of Tat peptides on DOPC and DOPG lipid bilayer membrane studied by molecular dynamics simulations[J].Molecular Simulation,2012,38(5):366-368.
[31]Kim,K.,Jordan,K..Particle mesh Ewald:An N[center-dot]log(N)method for Ewald sums in large systems[J].J.Chem.Phys,1994,98:10089.
[32]Hess,B.,Bekker,H.,Berendsen,H.J.,Fraaije,J.G..LINCS:a linear constraint solver for molecular simulations[J].Journal of computational chemistry,1997,18(12):1463-1472.
[33]张继伟,卞富永,施国军,徐四川.多巴胺在POPC磷脂双层膜中扩散和透过过程的分子动力学模拟[J].Acta Phys.-Chim.Sin,2014:30.
[34]张贇.人胰岛淀粉样多肽hIAPP单体及二聚体与POPG磷脂膜相互作用的分子动力学模拟研究[D].上海:复旦大学,2012.
[35]常钟文.淀粉样Aβ25-35片段寡聚体与POPG双层膜相互作用的分子动力学模拟[D].上海:复旦大学,2010.
[36]Fimland,G.,Eijsink,V.G.,Nissen-Meyer,J..Mutational analysis of the role of tryptophan residues in an antimicrobial peptide[J].Biochemistry,2002,41(30):9508-9515.
[37]Derossi,D.,Chassaing,G.,Prochiantz,A..Trojan peptides:the penetratin system for intracellular delivery[J].Trends in cell biology,1998,8(2):84-87.
[38]Richard,J.P.,Melikov,K.,Vives,E.,Ramos,C.,Verbeure,B.,Gait,M.J..Chernomordik,L.V.,Lebleu,B..Cell-penetrating peptides A reevaluation of the mechanism of cellular uptake[J].Journal of Biological Chemistry,2003,278(1):585-590.