Thermodynamic and Structural Analysis of Phosphotyrosine Polypeptide Binding to Grb2-SH2

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• 作者：Charles McNemar ; Mark E. Snow ; William T. Windsor ; Andrew Prongay ; Philip Mui ; Rumin Zhang ; James Durkin ; Hung V. Le ; and Patricia C. Weber
• 刊名：Biochemistry
• 出版年：1997
• 出版时间：August 19, 1997
• 年：1997
• 卷：36
• 期：33
• 页码：10006 - 10014
• 全文大小：216K
• 年卷期：v.36,no.33(August 19, 1997)
• ISSN：1520-4995

文摘

A thermodynamic analysis using isothermal titration calorimetry(ITC) has been performed toexamine the binding interaction between the SH2 (Srchomology 2) domain of growth factor receptorbinding protein 2 (Grb2-SH2) and one of its phosphotyrosine (pY)polypeptide ligands. Interaction ofthe Shc-derived phosphotyrosine hexapeptide Ac-SpYVNVQ-NH₂with Grb2-SH2 was both enthalpicallyand entropically favorable (H = -7.55 kcalmol^-1, -TS =-1.46 kcal mol^-1, G =-9.01 kcalmol^-1, T = 20 C). ITCexperiments using five alanine-substituted peptides were performed toexaminethe role of each side chain in binding. The results wereconsistent with homology models of the Grb2-SH2-Shc hexapeptide complex which identified several possible hydrogenbonds between Grb2-SH2and the phosphotyrosine and conserved asparagine(+2) side chainsof the Shc hexapeptide. These studiesalso demonstrated that the hydrophobic valine(+1) side chaincontributes significantly to the favorableentropic component of binding. The thermodynamic and structuraldata are consistent with a Grb2-SH2recognition motif of pY-hydrophobic-N-X (where X is any amino acidresidue). The measured heatcapacity of binding (C_p = -146cal mol^-1 K^-1) wasvery similar to computed values using semiempiricalestimates (C_p = -106 to -193cal mol^-1 K^-1) derived from apolar andpolar accessible surface areavalues calculated from several homology models of the Grb2-SH2-Shchexapeptide complex. Thehomology model which most closely reproduced the measuredC_p value is also the model whichhadthe lowest RMS deviation from the subsequently determined crystalstructure. Calculations based on thethermodynamic data and these semiempirical estimates indicated that thebinding event involves burial ofnearly comparable apolar (677 Ų) and polar (609Ų) surface areas.