Transition State Analogue Structures of Human Phosphoglycerate Kinase Establish the Importance of Charge Balance in Catalysis

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• 作者：Matthew J. Cliff ; Matthew W. Bowler ; Andrea Varga ; James P. Marston ; Judit Szab ; Andrea M. Hounslow ; Nicola J. Baxter ; G. Michael Blackburn ; Mria Vas ; Jonathan P. Waltho
• 刊名：Journal of the American Chemical Society
• 出版年：2010
• 出版时间：May 12, 2010
• 年：2010
• 卷：132
• 期：18
• 页码：6507-6516
• 全文大小：489K
• 年卷期：v.132,no.18(May 12, 2010)
• ISSN：1520-5126

文摘

Transition state analogue (TSA) complexes formed by phosphoglycerate kinase (PGK) have been used to test the hypothesis that balancing of charge within the transition state dominates enzyme-catalyzed phosphoryl transfer. High-resolution structures of trifluoromagnesate (MgF₃⁻) and tetrafluoroaluminate (AlF₄⁻) complexes of PGK have been determined using X-ray crystallography and ¹⁹F-based NMR methods, revealing the nature of the catalytically relevant state of this archetypal metabolic kinase. Importantly, the side chain of K219, which coordinates the α-phosphate group in previous ground state structures, is sequestered into coordinating the metal fluoride, thereby creating a charge environment complementary to the transferring phosphoryl group. In line with the dominance of charge balance in transition state organization, the substitution K219A induces a corresponding reduction in charge in the bound aluminum fluoride species, which changes to a trifluoroaluminate (AlF₃⁰) complex. The AlF₃⁰ moiety retains the octahedral geometry observed within AlF₄⁻ TSA complexes, which endorses the proposal that some of the widely reported trigonal AlF₃⁰ complexes of phosphoryl transfer enzymes may have been misassigned and in reality contain MgF₃⁻.