The 1.9 Å Resolution Crystal Structure of Phosphono-CheY, an Analogue of the Active Form of the Response Regulator, CheY

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• 作者：Christopher J. Halkides ; Megan M. McEvoy ; Eric Casper ; Philip Matsumura ; Karl Volz ; and Frederick W. Dahlquist
• 刊名：Biochemistry
• 出版年：2000
• 出版时间：May 9, 2000
• 年：2000
• 卷：39
• 期：18
• 页码：5280 - 5286
• 全文大小：274K
• 年卷期：v.39,no.18(May 9, 2000)
• ISSN：1520-4995

文摘

To structurally characterize the activated state of the transiently phosphorylated signaltransduction protein CheY, we have constructed an -thiophosphonate derivative of the CheY D57Cpoint mutant and determined its three-dimensional structure at 1.85 Å resolution. We have also characterizedthis analogue with high-resolution NMR and studied its binding to a peptide derived from FliM, CheY'starget component of the flagellar motor. The chemically modified derivative, phosphono-CheY, exhibitsmany of the chemical properties of phosphorylated wild-type CheY, except that it is indefinitely stable.Electron density for the -thiophosphonate substitution is clear and readily interpretable; omit refinementdensity at the phosphorus atom is greater than 10. The molecule shows a number of localizedconformational changes that are believed to constitute the postphosphorylation activation events. Themost obvious of these changes include movement of the side chain of the active site base, Lys 109, anda predominately buried conformation of the side chain of Tyr 106. In addition, there are a number ofmore subtle changes more distant from the active site involving the 4 and 5 helices. These results areconsistent with our previous structural interpretations of other CheY activation mutants, and with ourearlier hypotheses concerning CheY activation through propagation of structural changes away from theactive site.