BAM8-22的NMR溶液结构测定与BAM8-22及其片段肽的分子动力学研究
|
摘要
目的:对牛肾上腺髓质肽8-22(BAM8-22:Val-Gly-Arg-Pro-Glu-Trp-Trp-Met-Asp-Tyr-Gln-Lys-Arg-Tyr-Gly)进行首次空间结构的研究,并进一步结合已有的生物活性数据从三维结构水平上分析BAM8-22及其一系列片段肽的构效关系。方法:首先通过二维核磁共振技术测定了BAM8-22在水溶液中的空间三维构象,然后再通过20 ns的水中分子动力学模拟补充观察BAM8-22的动力学特征;接着,在BAM8-22核磁结构的基础上,对其片段肽BAM13-22,BAM15-22,BAM8-18,BAM8-20分别进行20 ns的水中分子动力学模拟,分析观察它们的结构变化与稳定性。结果:水溶液中的BAM8-22在肽段BAM12-20是一个相对稳定的α-螺旋结构,两端氨基酸残基是一个比较柔性的构象;而其片段肽BAM13-22的α-螺旋大约稳定在BAM15-20;BAM15-22的α-螺旋完全消失;BAM8-18的α-螺旋大约稳定在BAM12-16;BAM8-20的α-螺旋大约稳定在BAM12-18。结论:通过得到的BAM8-22的溶液三维结构与动力学特征及其片段肽的空间构象的变化信息,再结合已有的功能活性数据进行构效关系分析发现BAM12-20位的α-螺旋对维持BAM8-22的生物活性是非常必需的。所有这些结构研究,不仅能够指导合理改造BAM8-22,进而设计活性更好的类似物,同时也能给进一步研究BAM8-22与Mrg受体的相互作用机制提供结构基础。
Objective: To shed first light on the structural information of Bovine adrenal medullary peptide BAM8-22 (Val-Gly-Arg-Pro-Glu-Trp-Trp-Met-Asp-Tyr-Gln-Lys-Arg-Tyr- Gly), and to gain more detail insight into conformation-activity relationship of BAM8-22 and its truncated peptides on the basis of available function activity study. Methods: The solution structure of BAM8-22 was determined by 2D NMR in water. The dynamic features of BAM8-22 were further investigated by molecular dynamics simulation for 20 ns. Furthermore, the solution structure of BAM8-22 was used to study the conformation characterization of truncated peptide BAM13-22, BAM15-22, BAM8-18 and BAM8-20 by molecular dynamics simulation for 20 ns.
Results: BAM8-22 possesses a relatively well-definedα-helix structure during approximately BAM12-20 in aqueous solution, whereas the both termini show highly flexibility. Theα-helix structure of BAM13-22, BAM8-18 and BAM8-20 were spanning from BAM15-20, BAM12-16 and BAM12-18, respectively, while theα-helix structure in BAM15-22 was completely disappeared. Conclusion: From the three-dimentional structure and dynamics features of BAM8-22 and conformational characterization of its truncated peptides, combined with the available functional activity data, it seems reasonable to conclude that the well-definedα-helix structure plays an essential role on the bioactivcity of BAM8-22. All the biophysical data obtained in the current work provides important structural fundation of BAM8-22 which would be helpful to rationally design its analogues and also could be uesd as structural model for further studying its interaction with Mrg receptors.
Objective: To shed first light on the structural information of Bovine adrenal medullary peptide BAM8-22 (Val-Gly-Arg-Pro-Glu-Trp-Trp-Met-Asp-Tyr-Gln-Lys-Arg-Tyr- Gly), and to gain more detail insight into conformation-activity relationship of BAM8-22 and its truncated peptides on the basis of available function activity study. Methods: The solution structure of BAM8-22 was determined by 2D NMR in water. The dynamic features of BAM8-22 were further investigated by molecular dynamics simulation for 20 ns. Furthermore, the solution structure of BAM8-22 was used to study the conformation characterization of truncated peptide BAM13-22, BAM15-22, BAM8-18 and BAM8-20 by molecular dynamics simulation for 20 ns.
Results: BAM8-22 possesses a relatively well-definedα-helix structure during approximately BAM12-20 in aqueous solution, whereas the both termini show highly flexibility. Theα-helix structure of BAM13-22, BAM8-18 and BAM8-20 were spanning from BAM15-20, BAM12-16 and BAM12-18, respectively, while theα-helix structure in BAM15-22 was completely disappeared. Conclusion: From the three-dimentional structure and dynamics features of BAM8-22 and conformational characterization of its truncated peptides, combined with the available functional activity data, it seems reasonable to conclude that the well-definedα-helix structure plays an essential role on the bioactivcity of BAM8-22. All the biophysical data obtained in the current work provides important structural fundation of BAM8-22 which would be helpful to rationally design its analogues and also could be uesd as structural model for further studying its interaction with Mrg receptors.
引文
[1] Dong X, Han S, Zylka MJ, Simon MI, Anderson DJ. A diverse family of GPCRs expressed in specific subsets of nociceptive sensory neurons. Cell 2001;106:619-32.
[2] Lembo PM, Grazzini E, Groblewski T, O'Donnell D, Roy MO, Zhang J, et al. Proenkephalin A gene products activate a new family of sensory neuron-specific GPCRs. Nat Neurosci 2002;5:201-9.
[3] Zylka MJ, Dong XZ, Southwell AL, Anderson DJ. Atypical expansion in mice of the sensory neuronspecific Mrg G protein-coupled receptor family. Proceedings of the National Academy of Sciences of the United States of America 2003;100:10043-8.
[4] Grazzini E, Puma C, Roy MO, Yu XH, O'Donnell D, Schmidt R, et al. Sensory central neuron-specific receptor activation elicits and peripheral nociceptive effects in rats. Proceedings of the National Academy of Sciences of the United States of America 2004;101:7175-80.
[5] Lefkowitz RJ. The superfamily of heptahelical receptors. Nature Cell Biology, 2000. p. E133-E6.
[6] Kristiansen K. Molecular mechanisms of ligand binding, signaling, and regulation within the superfamily of G-protein-coupled receptors: molecular modeling and mutagenesis approaches to receptor structure and function. Pharmacology & Therapeutics 2004; 103:21-80.
[7] Evers A, Klabunde T. Structure-based drug discovery using GPCR homology modeling: successful virtual screening for antagonists of the alpha1A adrenergic receptor. J Med Chem 2005;48:1088-97.
[8] Bockaert J, Pin JP. Molecular tinkering of G protein-coupled receptors: an evolutionary success. EMBO J 1999;18:1723-9.
[9] Gether U. Uncovering molecular mechanisms involved in activation of G protein-coupled receptors. Endocr Rev 2000;21:90-113.
[10] Klabunde T, Hessler G. Drug design strategies for targeting G-protein-coupled receptors. Chembiochem 2002;3:928-44.
[11] Choi SS, Lahn BT. Adaptive evolution of MRG, a neuron-specific gene family implicated in nociception. Genome Research 2003; 13:2252-9.
[12] Simonin F, Kieffer BL. Two faces for an opioid peptide--and more receptors for pain research. Nat Neurosci 2002;5:185-6.
[13] Han SK, Dong X, Hwang JI, Zylka MJ, Anderson DJ, Simon MI. Orphan G protein-coupled receptors MrgA1 and MrgC11 are distinctively activated by RF-amide-related peptides through the Galpha q/11 pathway. Proc Natl Acad Sci U S A 2002;99:14740-5.
[14] Robas N, Mead E, Fidock M. MrgX2 is a high potency cortistatin receptor expressed in dorsal root ganglion. Journal of Biological Chemistry 2003;278:44400-4.
[15] Bender E, Buist A, Jurzak M, Langlois X, Baggerman G, Verhasselt P, et al. Characterization of an orphan G protein-coupled receptor localized in the dorsal root ganglia reveals adenine as a signaling molecule. Proc Natl Acad Sci U S A 2002;99:8573-8.
[16] Shinohara T, Harada M, Ogi K, Maruyama M, Fujii R, Tanaka H, et al. Identification of a G protein-coupled receptor specifically responsive to beta-alanine. J Biol Chem 2004;279:23559-64.
[17] Kamohara M, Matsuo A, Takasaki J, Kohda M, Matsumoto M, Matsumoto S, et al. Identification of MrgX2 as a human G-protein-coupled receptor for proadrenomedullin N-terminal peptides. Biochem Biophys Res Commun 2005;330:1146-52.
[18] Wang Z, Takahashi T, Saito Y, Nagasaki H, Ly NK, Nothacker HP, et al. Salusin beta is a surrogate ligand of the mas-like G protein-coupled receptor MrgA1. Eur J Pharmacol 2006;539:145-50.
[19] Kunapuli P, Lee S, Zheng W, Alberts M, Kornienko O, Mull R, et al. Identification of small molecule antagonists of the human mas-related gene-X1 receptor. Anal Biochem 2006;351:50-61.
[20] Malik L, Kelly NM, Ma JN, Currier EA, Burstein ES, Olsson R. Discovery of non-peptidergic MrgX1 and MrgX2 receptor agonists and exploration of an initial SAR using solid-phase synthesis. Bioorg Med Chem Lett 2009; 19:1729-32.
[21] Dray A, Nunan L, Wire W. Proenkephalin A fragments exhibit spinal and supraspinal opioid activity in vivo. J Pharmacol Exp Ther 1985;235:670-6.
[22] Hollt V, Haarmann I, Grimm C, Herz A, Tulunay FC, Loh HH. Pro-enkephalin intermediates in bovine brain and adrenal medulla: characterization of immunoreactive peptides related to BAM-22P and peptide F. Life Sci 1982;31:1883-6.
[23] Hollt V, Tulunay FC, Woo SK, Loh HH, Herz A. Opioid peptides derived from pro-enkephalin A but not that from pro-enkephalin B are substantial analgesics after administration into brain of mice. Eur J Pharmacol 1982;85:355-6.
[24] Mizuno K, Minamino N, Kangawa K, Matsuo H. A new family of endogenous "big" Met-enkephalins from bovine adrenal medulla: purification and structure of docosa-(BAM-22P) and eicosapeptide (BAM-20P) with very potent opiate activity. Biochem Biophys Res Commun 1980;97:1283-90.
[25] Mizuno K, Minamino N, Kangawa K, Matsuo H. A new endogenous opioid peptide from bovine adrenal medulla: isolation and amino acid sequence of a dodecapeptide (BAM-12P). Biochem Biophys Res Commun 1980;95:1482-8.
[26] Swain MG, MacArthur L, Vergalla J, Jones EA. Adrenal secretion of BAM-22P, a potent opioid peptide, is enhanced in rats with acute cholestasis. Am J Physiol 1994;266:G201-5.
[27] Sanchez-Blazquez P, Garzon J. Opioid activity of pro-enkephalin-derived peptides in mouse vas deferens and guinea pig ileum. Neurosci Lett 1985;61:267-71.
[28] Wollemann M, Benyhe S. Non-opioid actions of opioid peptides. Life Sci 2004;75:257-70.
[29] Cai M, Chen T, Quirion R, Hong Y. The involvement of spinal bovine adrenal medulla 22-like peptide, the proenkephalin derivative, in modulation of nociceptive processing. Eur J Neurosci 2007;26:1128-38.
[30] Cox PJ, Pitcher T, Trim SA, Bell CH, Qin W, Kinloch RA. The effect of deletion of the orphan G - protein coupled receptor (GPCR) gene MrgE on pain-like behaviours in mice. Mol Pain 2008;4:2.
[31] Hong Y, Dai P, Jiang J, Zeng X. Dual effects of intrathecal BAM22 on nociceptive responses in acute and persistent pain-potential function of a novel receptor. Br J Pharmacol 2004; 141:423-30.
[32] Zeng X, Huang H, Hong Y. Effects of intrathecal BAM22 on noxious stimulus-evoked c-fos expression in the rat spinal dorsal horn.Brain Res 2004;1028:170-9.
[33]Chen H,Ikeda SR.Modulation of ion channels and synaptic transmission by a human sensory neuron-specific G-protein-coupled receptor,SNSR4/mrgX1,heterologously expressed in cultured rat neurons.J Neurosci 2004;24:5044-53.
[34]Jiang J,Huang J,Hong Y.Bovine adrenal medulla 22 reverses antinociceptive morphine tolerance in the rat.Behav Brain Res 2005;168:167-71.
[35]Hager UA,Hein A,Lennerz JK,Zimmermann K,Neuhuber WL,Reeh PW.Morphological characterization of rat Mas-related G-protein-coupled receptor C and functional analysis of agonists.Neuroscience 2008;151:242-54.
[36]Gustafson EL,Maguire M,Campanella M,Tarozzo G,Jia Y,Dong XW,et al.Regulation of two rat mas-related genes in a model of neuropathic pain.Brain Res Mol Brain Res 2005;142:58-64.
[37]Burstein ES,Ott TR,Feddock M,Ma JN,Fuhs S,Wong S,et al.Characterization of the Mas-related gene family:structural and functional conservation of human and rhesus MrgX receptors.British Journal of Pharmacology 2006;147:73-82.
[38]Breit A,Gagnidze K,Devi LA,Lagace M,Bouvier M.Simultaneous activation of the delta opioid receptor(deltaOR)/sensory neuron-specific receptor-4(SNSR-4) hetero-oligomer by the mixed bivalent agonist bovine adrenal medulla peptide 22 activates SNSR-4 but inhibits deltaOR signaling.Mol Pharmacol 2006;70:686-96.
[39]Cai Q,Jiang J,Chen T,Hong Y.Sensory neuron-specific receptor agonist BAM8-22 inhibits the development and expression of tolerance to morphine in rats.Behav Brain Res 2007;178:154-9.
[40]Chen P,Liu Y,Hong Y.Effect of chronic administration of morphine on the expression of bovine adrenal medulla 22-like immunoreactivity in the spinal cord of rats.Eur J Pharmacol 2008;589:110-3.
[41]Chen T,Hu Z,Quirion R,Hong Y.Modulation of NMDA receptors by intrathecal administration of the sensory neuron-specific receptor agonist BAM8-22.Neuropharmacology 2008;54:796-803.
[42]Young JK,Anklin C,Hicks RP.NMR and molecular modeling investigations of the neuropeptide substance P in the presence of 15 mM sodium dodecyl sulfate micelles.Biopolymers 1994;34:1449-62.
[43]Young JK,Hicks RP.Nmr and molecular modeling investigations of the neuropeptide bradykinin in three different solvent systems:DMSO,9:1 dioxane/water,and in the presence of 7.4 mM lyso phosphatidylcholine micelles.Biopolymers 1994;34:611-23.
[44]Anglister J,Scherf T,Zilber B,Levy R.Two-dimensional NMR studies of the interactions between a peptide of cholera toxin and monoclonal antibodies.Biopolymers 1995;37:383-9.
[45]夏佑林,吴季辉,刘琴,施蕴渝.生物大分子多维核磁共振.中国科学技术大学出社1999:118-49.
[46]阎隆飞.蛋白质分子结构.清华大学出版社 1999.
[47]Wuthrich K,Wider G,Wagner G,Braun W.Sequential resonance assignments as a basis for determination of spatial protein structures by high resolution proton nuclear magnetic resonance.J Mol Biol 1982;155:311-9.
[48]Billeter M,Braun W,Wuthrich K.Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra. Computation of sterically allowed proton-proton distances and statistical analysis of proton-proton distances in single crystal protein conformations. J Mol Biol 1982;155:321-46.
[49] Wagner G, Wuthrich K. Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra. Basic pancreatic trypsin inhibitor. J Mol Biol 1982;155:347-66.
[50] Wider G, Lee KH, Wuthrich K. Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra. Glucagon bound to perdeuterated dodecylphosphocholine micelles. J Mol Biol 1982; 155:367-88.
[51] Havel TFaW, K., . A distance geometry program for determining the structures of small proteins and other macromolecules from nuclear magnetic resonance measurements of intramolecular HH proximities in solution. Bull Math Bio 1984; 46:673-98.
[52] Kaptein R, Zuiderweg ER, Scheek RM, Boelens R, van Gunsteren WF. A protein structure from nuclear magnetic resonance data. lac repressor headpiece. J Mol Biol 1985; 182:179-82.
[53] Wuthrich. K. NMR of Proteins and Nucleic Acids. John Wiley: New York 1986.
[54] Pardi A, Billeter M, Wuthrich K. Calibration of the angular dependence of the amide proton-C alpha proton coupling constants, 3JHN alpha, in a globular protein. Use of 3JHN alpha for identification of helical secondary structure. J Mol Biol 1984;180:741-51.
[55] Dyson HJ, Wright PE. Defining solution conformations of small linear peptides. Annu Rev Biophys Biophys Chem 1991;20:519-38.
[56] Wishart DS, Sykes BD, Richards FM. Relationship between nuclear magnetic resonance chemical shift and protein secondary structure. J Mol Biol 1991;222:311-33.
[57] Wishart DS, Sykes BD, Richards FM. The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy. Biochemistry 1992;31:1647-51.
[58] Wishart DS, Sykes BD, Richards FM. Simple techniques for the quantification of protein secondary structure by 1H NMR spectroscopy. FEBS Lett 1991;293:72-80.
[59] Wishart DS, Sykes BD. The 13C chemical-shift index: a simple method for the identification of protein secondary structure using 13C chemical-shift data. J Biomol NMR 1994;4:171-80.
[60] Keepers JWaJ, T. L. A Theoretical Study of Distance Determinations from NMR. Two-Dimensional Nuclear Overhauser Effect Spectra. J Magn Reson 1984;57:404~26.
[61] Brunger AT, Adams PD, Clore GM, DeLano WL, Gros P, Grosse-Kunstleve RW, et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr D Biol Crystallogr 1998;54:905-21.
[62] Brunger AT. Version 1.2 of the Crystallography and NMR system. Nat Protoc 2007;2:2728-33.
[63] Laskowski RA, Rullmannn JA, MacArthur MW, Kaptein R, Thornton JM. AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR. J Biomol NMR 1996;8:477-86.
[64] Cavanagh J, Fairbrother, W.J., Palmer III, A.G., Skelton, N.J. Protein NMR Spectroscopy: Principles and Practice. 1996.
[65] Wuthrich K. The way to NMR structures of proteins. Nature Structural Biology 2001 ;8:923-5.
[66] Trzesniak Da, Glattli, A.a , Jaun, B.b , Van Gunsteren, W.F. Interpreting NMR data for P-peptides using molecular dynamics simulations. J Am Chem Soc 2005;127:14320-9
[67] Karplus M, McCammon, J.A. Molecular dynamics simulations of biomolecules. Nature Structural Biology 2002;9:646-52
[68]Karplus M.Molecular dynamics simulations of biomolecules.Accounts of Chemical Research 2002 35:321-3
[69]Karplus MK,J.Molecular dynamics and protein function.Proceedings of the National Academy of Sciences of the United States of America 2005;102:6679-85
[70]Norberg J,Nilsson,L.Advances in biomolecular simulations:Methodology and recent applications.Quarterly Reviews of Biophysics 2003 36:257-306
[71]Zagrovic B,Van Gunsteren,W.F.Comparing atomistic simulation data with the NMR experiment:How much can NOEs actually tell us? Proteins:Structure,Function and Genetics 2006;63:210-8.
[72]Faramarz Mehrnejad HN-M,Bijan Ranjbar The structural properties of magainin in water,TFE/water,and aqueous urea solutions:Molecular dynamics simulations Proteins:Structure,Function,and Bioinformatics 2007;67:931-40.
[73]Van Der Spoel D,Lindahl E,Hess B,Groenhof G,Mark AE,Berendsen HJ.GROMACS:fast,flexible,and free.J Comput Chem 2005;26:1701-18.
[74]Pearlman DA,Case DA,Caldwell JW,Ross WS,Cheatham TE,Debolt S,et al.AMBER,a package of computer programs for applying molecular mechanics,normal mode analysis,molecular dynamics and free energy calculations to simulate the structural and energetic properties of molecules.Comput Phys Commun 1995;91:1-41.
[75]Brooks BR,Bruccoleri R,Olafson BD,States DJ,Swaminathan S,Karplus M.CHARMM:A program for macromolecular energy,minmimization,and dynamics calculations4 J Comp Chem 1983;4:187-217.
[76]Phillips JC,Braun R,Wang W,Gumbart J,Tajkhorshid E,Villa E,et al.Scalable molecular dynamics with NAMD.J Comput Chem 2005;26:1781-802.
[77]Brooks BR,Bruccoleri,R.E.,Olafson,B.D.,States,D.J.,Swaminathan,S.,Karplus,M..CHARMM:A program for macromolecular energy,minmimization,and dynamics calculations.J Comp Chem 1983;4.
[78]MacKerell AD,Bashford D,Bellott M,Dunbrack RL,Evanseck JD,Field MJ,et al.All-atom empirical potential for molecular modeling and dynamics studies of proteins.J Phys Chem B 1998;102:3586-616.
[79]Mackerell AD,Wiorkiewiczkuczera J,Karplus M.An all-atom empirical energy function for the simulation of nucleic acids.J Am Chem Soc 1995;117:11946-75.
[80]Berendsen WFvGaHJC.Groningen Molecular Simulation(GROMOS) Library Manual.Biomos 1987.
[81]Oostenbrink C,Villa A,Mark AE,van Gunsteren WF.A biomolecular force field based on the free enthalpy of hydration and solvation:the GROMOS force-field parameter sets 53A5 and 53A6.J Comput Chem 2004;25:1656-76.
[82]Jorgensen WL,Maxwell,D.S,Tirado-Rives,J.Development and testing of the OPLS all-atom force field on conformational energetics and properties of organic liquids.J Am Chem Soc 1996;118:11225-36
[83]Jorgensen WL,Tirado-Rives,J.The OPLS potential functions for proteins.Energy minimizations for crystals of cyclic peptides and crambin.J Am Chem Soc 1988;110:1657-66.
[84]Dauber-Osguthorpe PR,V.A.;Osguthorpe,D.J.;Wolff,J.;Genest,M.;Hagler,A.T.Structure and energetics of ligand binding to proteins:E.coli dihydrofolate reductase-trimethoprim,a drug-receptor system.Proteins:Structure,Function and Genetics 1988;4:31-47.
[85]DeLano WL.The case for open-source software in drug discovery.Drug Discov Today 2005;10:213-7.
[86]Berendsen HJC,van der Spoel D,van Drunen R.GROMACS:A message-passing parallel molecular dynamics implementation.Computer Physics Communications 1995;91:43-56.
[87]Kabsch W,Sander C.Dictionary of protein secondary structure:pattern recognition of hydrogen-bonded and geometrical features.Biopolymers 1983;22:2577-637.
[88]W(u¨)thrich K.NMR of Proteins and Nucleic Acids.John Wiley:New York 1986.
[89]Alonso H,Bliznyuk AA,Gready JE.Combining docking and molecular dynamic simulations in drug design.Med Res Rev 2006;26:531-68.
[90]Carlson HA,McCammon JA.Accommodating protein flexibility in computational drug design.Molecular Pharmacology 2000;57:213-8.
[91]Verkhivker GM,Bouzida D,Gehlhaar DK,Rejto PA,Freer ST,Rose PW.Complexity and simplicity of ligand-macromolecule interactions:the energy landscape perspective.Curr Opin Struct Biol 2002;12:197-203.
[92]Glattli A,van Gunsteren WF.Are NMR-derived model structures for beta-peptides representative for the ensemble of structures adopted in solution? Angew Chem Int Ed Engl 2004;43:6312-6.
[93]Heo JY,Han SK,Vaidehi N,Wendel J,Kekenes-Huskey P,Goddard WA.Prediction of the 3D structure of FMRF-amide neuropeptides bound to the mouse MrgC11 GPCR and experimental validation.Chembiochem 2007;8:1527-39.
[94]Martinelli A,Tuccinardi T.Molecular modeling of adenosine receptors:new results and trends.Med Res Rev 2008;28:247-77.
[95]Patny A,Desai PV,Avery MA.Homology modeling of G-protein-coupled receptors and implications in drug design.Curr Med Chem 2006;13:1667-91.
[2] Lembo PM, Grazzini E, Groblewski T, O'Donnell D, Roy MO, Zhang J, et al. Proenkephalin A gene products activate a new family of sensory neuron-specific GPCRs. Nat Neurosci 2002;5:201-9.
[3] Zylka MJ, Dong XZ, Southwell AL, Anderson DJ. Atypical expansion in mice of the sensory neuronspecific Mrg G protein-coupled receptor family. Proceedings of the National Academy of Sciences of the United States of America 2003;100:10043-8.
[4] Grazzini E, Puma C, Roy MO, Yu XH, O'Donnell D, Schmidt R, et al. Sensory central neuron-specific receptor activation elicits and peripheral nociceptive effects in rats. Proceedings of the National Academy of Sciences of the United States of America 2004;101:7175-80.
[5] Lefkowitz RJ. The superfamily of heptahelical receptors. Nature Cell Biology, 2000. p. E133-E6.
[6] Kristiansen K. Molecular mechanisms of ligand binding, signaling, and regulation within the superfamily of G-protein-coupled receptors: molecular modeling and mutagenesis approaches to receptor structure and function. Pharmacology & Therapeutics 2004; 103:21-80.
[7] Evers A, Klabunde T. Structure-based drug discovery using GPCR homology modeling: successful virtual screening for antagonists of the alpha1A adrenergic receptor. J Med Chem 2005;48:1088-97.
[8] Bockaert J, Pin JP. Molecular tinkering of G protein-coupled receptors: an evolutionary success. EMBO J 1999;18:1723-9.
[9] Gether U. Uncovering molecular mechanisms involved in activation of G protein-coupled receptors. Endocr Rev 2000;21:90-113.
[10] Klabunde T, Hessler G. Drug design strategies for targeting G-protein-coupled receptors. Chembiochem 2002;3:928-44.
[11] Choi SS, Lahn BT. Adaptive evolution of MRG, a neuron-specific gene family implicated in nociception. Genome Research 2003; 13:2252-9.
[12] Simonin F, Kieffer BL. Two faces for an opioid peptide--and more receptors for pain research. Nat Neurosci 2002;5:185-6.
[13] Han SK, Dong X, Hwang JI, Zylka MJ, Anderson DJ, Simon MI. Orphan G protein-coupled receptors MrgA1 and MrgC11 are distinctively activated by RF-amide-related peptides through the Galpha q/11 pathway. Proc Natl Acad Sci U S A 2002;99:14740-5.
[14] Robas N, Mead E, Fidock M. MrgX2 is a high potency cortistatin receptor expressed in dorsal root ganglion. Journal of Biological Chemistry 2003;278:44400-4.
[15] Bender E, Buist A, Jurzak M, Langlois X, Baggerman G, Verhasselt P, et al. Characterization of an orphan G protein-coupled receptor localized in the dorsal root ganglia reveals adenine as a signaling molecule. Proc Natl Acad Sci U S A 2002;99:8573-8.
[16] Shinohara T, Harada M, Ogi K, Maruyama M, Fujii R, Tanaka H, et al. Identification of a G protein-coupled receptor specifically responsive to beta-alanine. J Biol Chem 2004;279:23559-64.
[17] Kamohara M, Matsuo A, Takasaki J, Kohda M, Matsumoto M, Matsumoto S, et al. Identification of MrgX2 as a human G-protein-coupled receptor for proadrenomedullin N-terminal peptides. Biochem Biophys Res Commun 2005;330:1146-52.
[18] Wang Z, Takahashi T, Saito Y, Nagasaki H, Ly NK, Nothacker HP, et al. Salusin beta is a surrogate ligand of the mas-like G protein-coupled receptor MrgA1. Eur J Pharmacol 2006;539:145-50.
[19] Kunapuli P, Lee S, Zheng W, Alberts M, Kornienko O, Mull R, et al. Identification of small molecule antagonists of the human mas-related gene-X1 receptor. Anal Biochem 2006;351:50-61.
[20] Malik L, Kelly NM, Ma JN, Currier EA, Burstein ES, Olsson R. Discovery of non-peptidergic MrgX1 and MrgX2 receptor agonists and exploration of an initial SAR using solid-phase synthesis. Bioorg Med Chem Lett 2009; 19:1729-32.
[21] Dray A, Nunan L, Wire W. Proenkephalin A fragments exhibit spinal and supraspinal opioid activity in vivo. J Pharmacol Exp Ther 1985;235:670-6.
[22] Hollt V, Haarmann I, Grimm C, Herz A, Tulunay FC, Loh HH. Pro-enkephalin intermediates in bovine brain and adrenal medulla: characterization of immunoreactive peptides related to BAM-22P and peptide F. Life Sci 1982;31:1883-6.
[23] Hollt V, Tulunay FC, Woo SK, Loh HH, Herz A. Opioid peptides derived from pro-enkephalin A but not that from pro-enkephalin B are substantial analgesics after administration into brain of mice. Eur J Pharmacol 1982;85:355-6.
[24] Mizuno K, Minamino N, Kangawa K, Matsuo H. A new family of endogenous "big" Met-enkephalins from bovine adrenal medulla: purification and structure of docosa-(BAM-22P) and eicosapeptide (BAM-20P) with very potent opiate activity. Biochem Biophys Res Commun 1980;97:1283-90.
[25] Mizuno K, Minamino N, Kangawa K, Matsuo H. A new endogenous opioid peptide from bovine adrenal medulla: isolation and amino acid sequence of a dodecapeptide (BAM-12P). Biochem Biophys Res Commun 1980;95:1482-8.
[26] Swain MG, MacArthur L, Vergalla J, Jones EA. Adrenal secretion of BAM-22P, a potent opioid peptide, is enhanced in rats with acute cholestasis. Am J Physiol 1994;266:G201-5.
[27] Sanchez-Blazquez P, Garzon J. Opioid activity of pro-enkephalin-derived peptides in mouse vas deferens and guinea pig ileum. Neurosci Lett 1985;61:267-71.
[28] Wollemann M, Benyhe S. Non-opioid actions of opioid peptides. Life Sci 2004;75:257-70.
[29] Cai M, Chen T, Quirion R, Hong Y. The involvement of spinal bovine adrenal medulla 22-like peptide, the proenkephalin derivative, in modulation of nociceptive processing. Eur J Neurosci 2007;26:1128-38.
[30] Cox PJ, Pitcher T, Trim SA, Bell CH, Qin W, Kinloch RA. The effect of deletion of the orphan G - protein coupled receptor (GPCR) gene MrgE on pain-like behaviours in mice. Mol Pain 2008;4:2.
[31] Hong Y, Dai P, Jiang J, Zeng X. Dual effects of intrathecal BAM22 on nociceptive responses in acute and persistent pain-potential function of a novel receptor. Br J Pharmacol 2004; 141:423-30.
[32] Zeng X, Huang H, Hong Y. Effects of intrathecal BAM22 on noxious stimulus-evoked c-fos expression in the rat spinal dorsal horn.Brain Res 2004;1028:170-9.
[33]Chen H,Ikeda SR.Modulation of ion channels and synaptic transmission by a human sensory neuron-specific G-protein-coupled receptor,SNSR4/mrgX1,heterologously expressed in cultured rat neurons.J Neurosci 2004;24:5044-53.
[34]Jiang J,Huang J,Hong Y.Bovine adrenal medulla 22 reverses antinociceptive morphine tolerance in the rat.Behav Brain Res 2005;168:167-71.
[35]Hager UA,Hein A,Lennerz JK,Zimmermann K,Neuhuber WL,Reeh PW.Morphological characterization of rat Mas-related G-protein-coupled receptor C and functional analysis of agonists.Neuroscience 2008;151:242-54.
[36]Gustafson EL,Maguire M,Campanella M,Tarozzo G,Jia Y,Dong XW,et al.Regulation of two rat mas-related genes in a model of neuropathic pain.Brain Res Mol Brain Res 2005;142:58-64.
[37]Burstein ES,Ott TR,Feddock M,Ma JN,Fuhs S,Wong S,et al.Characterization of the Mas-related gene family:structural and functional conservation of human and rhesus MrgX receptors.British Journal of Pharmacology 2006;147:73-82.
[38]Breit A,Gagnidze K,Devi LA,Lagace M,Bouvier M.Simultaneous activation of the delta opioid receptor(deltaOR)/sensory neuron-specific receptor-4(SNSR-4) hetero-oligomer by the mixed bivalent agonist bovine adrenal medulla peptide 22 activates SNSR-4 but inhibits deltaOR signaling.Mol Pharmacol 2006;70:686-96.
[39]Cai Q,Jiang J,Chen T,Hong Y.Sensory neuron-specific receptor agonist BAM8-22 inhibits the development and expression of tolerance to morphine in rats.Behav Brain Res 2007;178:154-9.
[40]Chen P,Liu Y,Hong Y.Effect of chronic administration of morphine on the expression of bovine adrenal medulla 22-like immunoreactivity in the spinal cord of rats.Eur J Pharmacol 2008;589:110-3.
[41]Chen T,Hu Z,Quirion R,Hong Y.Modulation of NMDA receptors by intrathecal administration of the sensory neuron-specific receptor agonist BAM8-22.Neuropharmacology 2008;54:796-803.
[42]Young JK,Anklin C,Hicks RP.NMR and molecular modeling investigations of the neuropeptide substance P in the presence of 15 mM sodium dodecyl sulfate micelles.Biopolymers 1994;34:1449-62.
[43]Young JK,Hicks RP.Nmr and molecular modeling investigations of the neuropeptide bradykinin in three different solvent systems:DMSO,9:1 dioxane/water,and in the presence of 7.4 mM lyso phosphatidylcholine micelles.Biopolymers 1994;34:611-23.
[44]Anglister J,Scherf T,Zilber B,Levy R.Two-dimensional NMR studies of the interactions between a peptide of cholera toxin and monoclonal antibodies.Biopolymers 1995;37:383-9.
[45]夏佑林,吴季辉,刘琴,施蕴渝.生物大分子多维核磁共振.中国科学技术大学出社1999:118-49.
[46]阎隆飞.蛋白质分子结构.清华大学出版社 1999.
[47]Wuthrich K,Wider G,Wagner G,Braun W.Sequential resonance assignments as a basis for determination of spatial protein structures by high resolution proton nuclear magnetic resonance.J Mol Biol 1982;155:311-9.
[48]Billeter M,Braun W,Wuthrich K.Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra. Computation of sterically allowed proton-proton distances and statistical analysis of proton-proton distances in single crystal protein conformations. J Mol Biol 1982;155:321-46.
[49] Wagner G, Wuthrich K. Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra. Basic pancreatic trypsin inhibitor. J Mol Biol 1982;155:347-66.
[50] Wider G, Lee KH, Wuthrich K. Sequential resonance assignments in protein 1H nuclear magnetic resonance spectra. Glucagon bound to perdeuterated dodecylphosphocholine micelles. J Mol Biol 1982; 155:367-88.
[51] Havel TFaW, K., . A distance geometry program for determining the structures of small proteins and other macromolecules from nuclear magnetic resonance measurements of intramolecular HH proximities in solution. Bull Math Bio 1984; 46:673-98.
[52] Kaptein R, Zuiderweg ER, Scheek RM, Boelens R, van Gunsteren WF. A protein structure from nuclear magnetic resonance data. lac repressor headpiece. J Mol Biol 1985; 182:179-82.
[53] Wuthrich. K. NMR of Proteins and Nucleic Acids. John Wiley: New York 1986.
[54] Pardi A, Billeter M, Wuthrich K. Calibration of the angular dependence of the amide proton-C alpha proton coupling constants, 3JHN alpha, in a globular protein. Use of 3JHN alpha for identification of helical secondary structure. J Mol Biol 1984;180:741-51.
[55] Dyson HJ, Wright PE. Defining solution conformations of small linear peptides. Annu Rev Biophys Biophys Chem 1991;20:519-38.
[56] Wishart DS, Sykes BD, Richards FM. Relationship between nuclear magnetic resonance chemical shift and protein secondary structure. J Mol Biol 1991;222:311-33.
[57] Wishart DS, Sykes BD, Richards FM. The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy. Biochemistry 1992;31:1647-51.
[58] Wishart DS, Sykes BD, Richards FM. Simple techniques for the quantification of protein secondary structure by 1H NMR spectroscopy. FEBS Lett 1991;293:72-80.
[59] Wishart DS, Sykes BD. The 13C chemical-shift index: a simple method for the identification of protein secondary structure using 13C chemical-shift data. J Biomol NMR 1994;4:171-80.
[60] Keepers JWaJ, T. L. A Theoretical Study of Distance Determinations from NMR. Two-Dimensional Nuclear Overhauser Effect Spectra. J Magn Reson 1984;57:404~26.
[61] Brunger AT, Adams PD, Clore GM, DeLano WL, Gros P, Grosse-Kunstleve RW, et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr D Biol Crystallogr 1998;54:905-21.
[62] Brunger AT. Version 1.2 of the Crystallography and NMR system. Nat Protoc 2007;2:2728-33.
[63] Laskowski RA, Rullmannn JA, MacArthur MW, Kaptein R, Thornton JM. AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR. J Biomol NMR 1996;8:477-86.
[64] Cavanagh J, Fairbrother, W.J., Palmer III, A.G., Skelton, N.J. Protein NMR Spectroscopy: Principles and Practice. 1996.
[65] Wuthrich K. The way to NMR structures of proteins. Nature Structural Biology 2001 ;8:923-5.
[66] Trzesniak Da, Glattli, A.a , Jaun, B.b , Van Gunsteren, W.F. Interpreting NMR data for P-peptides using molecular dynamics simulations. J Am Chem Soc 2005;127:14320-9
[67] Karplus M, McCammon, J.A. Molecular dynamics simulations of biomolecules. Nature Structural Biology 2002;9:646-52
[68]Karplus M.Molecular dynamics simulations of biomolecules.Accounts of Chemical Research 2002 35:321-3
[69]Karplus MK,J.Molecular dynamics and protein function.Proceedings of the National Academy of Sciences of the United States of America 2005;102:6679-85
[70]Norberg J,Nilsson,L.Advances in biomolecular simulations:Methodology and recent applications.Quarterly Reviews of Biophysics 2003 36:257-306
[71]Zagrovic B,Van Gunsteren,W.F.Comparing atomistic simulation data with the NMR experiment:How much can NOEs actually tell us? Proteins:Structure,Function and Genetics 2006;63:210-8.
[72]Faramarz Mehrnejad HN-M,Bijan Ranjbar The structural properties of magainin in water,TFE/water,and aqueous urea solutions:Molecular dynamics simulations Proteins:Structure,Function,and Bioinformatics 2007;67:931-40.
[73]Van Der Spoel D,Lindahl E,Hess B,Groenhof G,Mark AE,Berendsen HJ.GROMACS:fast,flexible,and free.J Comput Chem 2005;26:1701-18.
[74]Pearlman DA,Case DA,Caldwell JW,Ross WS,Cheatham TE,Debolt S,et al.AMBER,a package of computer programs for applying molecular mechanics,normal mode analysis,molecular dynamics and free energy calculations to simulate the structural and energetic properties of molecules.Comput Phys Commun 1995;91:1-41.
[75]Brooks BR,Bruccoleri R,Olafson BD,States DJ,Swaminathan S,Karplus M.CHARMM:A program for macromolecular energy,minmimization,and dynamics calculations4 J Comp Chem 1983;4:187-217.
[76]Phillips JC,Braun R,Wang W,Gumbart J,Tajkhorshid E,Villa E,et al.Scalable molecular dynamics with NAMD.J Comput Chem 2005;26:1781-802.
[77]Brooks BR,Bruccoleri,R.E.,Olafson,B.D.,States,D.J.,Swaminathan,S.,Karplus,M..CHARMM:A program for macromolecular energy,minmimization,and dynamics calculations.J Comp Chem 1983;4.
[78]MacKerell AD,Bashford D,Bellott M,Dunbrack RL,Evanseck JD,Field MJ,et al.All-atom empirical potential for molecular modeling and dynamics studies of proteins.J Phys Chem B 1998;102:3586-616.
[79]Mackerell AD,Wiorkiewiczkuczera J,Karplus M.An all-atom empirical energy function for the simulation of nucleic acids.J Am Chem Soc 1995;117:11946-75.
[80]Berendsen WFvGaHJC.Groningen Molecular Simulation(GROMOS) Library Manual.Biomos 1987.
[81]Oostenbrink C,Villa A,Mark AE,van Gunsteren WF.A biomolecular force field based on the free enthalpy of hydration and solvation:the GROMOS force-field parameter sets 53A5 and 53A6.J Comput Chem 2004;25:1656-76.
[82]Jorgensen WL,Maxwell,D.S,Tirado-Rives,J.Development and testing of the OPLS all-atom force field on conformational energetics and properties of organic liquids.J Am Chem Soc 1996;118:11225-36
[83]Jorgensen WL,Tirado-Rives,J.The OPLS potential functions for proteins.Energy minimizations for crystals of cyclic peptides and crambin.J Am Chem Soc 1988;110:1657-66.
[84]Dauber-Osguthorpe PR,V.A.;Osguthorpe,D.J.;Wolff,J.;Genest,M.;Hagler,A.T.Structure and energetics of ligand binding to proteins:E.coli dihydrofolate reductase-trimethoprim,a drug-receptor system.Proteins:Structure,Function and Genetics 1988;4:31-47.
[85]DeLano WL.The case for open-source software in drug discovery.Drug Discov Today 2005;10:213-7.
[86]Berendsen HJC,van der Spoel D,van Drunen R.GROMACS:A message-passing parallel molecular dynamics implementation.Computer Physics Communications 1995;91:43-56.
[87]Kabsch W,Sander C.Dictionary of protein secondary structure:pattern recognition of hydrogen-bonded and geometrical features.Biopolymers 1983;22:2577-637.
[88]W(u¨)thrich K.NMR of Proteins and Nucleic Acids.John Wiley:New York 1986.
[89]Alonso H,Bliznyuk AA,Gready JE.Combining docking and molecular dynamic simulations in drug design.Med Res Rev 2006;26:531-68.
[90]Carlson HA,McCammon JA.Accommodating protein flexibility in computational drug design.Molecular Pharmacology 2000;57:213-8.
[91]Verkhivker GM,Bouzida D,Gehlhaar DK,Rejto PA,Freer ST,Rose PW.Complexity and simplicity of ligand-macromolecule interactions:the energy landscape perspective.Curr Opin Struct Biol 2002;12:197-203.
[92]Glattli A,van Gunsteren WF.Are NMR-derived model structures for beta-peptides representative for the ensemble of structures adopted in solution? Angew Chem Int Ed Engl 2004;43:6312-6.
[93]Heo JY,Han SK,Vaidehi N,Wendel J,Kekenes-Huskey P,Goddard WA.Prediction of the 3D structure of FMRF-amide neuropeptides bound to the mouse MrgC11 GPCR and experimental validation.Chembiochem 2007;8:1527-39.
[94]Martinelli A,Tuccinardi T.Molecular modeling of adenosine receptors:new results and trends.Med Res Rev 2008;28:247-77.
[95]Patny A,Desai PV,Avery MA.Homology modeling of G-protein-coupled receptors and implications in drug design.Curr Med Chem 2006;13:1667-91.