PMID: 2496745Jan 24, 1989Paper

Probing the functional role of threonine-113 of Escherichia coli dihydrofolate reductase for its effect on turnover efficiency, catalysis, and binding

Biochemistry
C A Fierke, S J Benkovic

Abstract

The role of Thr-113 of Escherichia coli dihydrofolate reductase in binding and catalysis was probed by amino acid substitution. Thr-113, a strictly conserved residue that forms a hydrogen bond to the active-site Asp-27 and to the amino group of methotrexate through a fixed water molecule, was replaced by valine. The kinetic scheme is identical in form with the wild-type scheme, although many of the rate constants vary, including a decrease in the association rate constants and an increase in the dissociation rate constants for folate ligands, a decrease in the hydride-transfer rate constant in both directions, and an increase in the intrinsic pKa of Asp-27. Overall, replacement of Thr-113 by Val decreases the binding of folate substrates by approximately 2.3 kcal/mol. These multiple complex changes on various ground and transition states underscore the optimal properties of a strictly conserved residue in the evolution of catalytic function.

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Citations

Feb 1, 2006·Biochemistry·Lin WangAmnon Kohen
Sep 20, 2011·Chemical Reviews·Jeeyeon Lee, Nina M Goodey
Dec 17, 2011·Journal of the American Chemical Society·Vanja StojkovićAmnon Kohen
Oct 18, 2000·Proceedings of the National Academy of Sciences of the United States of America·H PanV J Hilser
Oct 13, 2006·Proceedings of the National Academy of Sciences of the United States of America·Lin WangAmnon Kohen
Dec 30, 2014·Accounts of Chemical Research·Gira BhabhaJames S Fraser
Jun 5, 2007·The FEBS Journal·Ronnie A BöckWilliam W Barrow
May 14, 1998·Chemistry & Biology·G P Miller, S J Benkovic
Jun 22, 2021·Annual Review of Biochemistry·Stephen J Benkovic

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