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Increased dynamic effects in a catalytically compromised variant of Escherichia coli dihydrofolate reductase

Ruiz-Pernia, J. Javier, Luk, Louis Yu Pan ORCID:, Garcia-Meseguer, Rafael, Martí, Sergio, Loveridge, Edric Joel, Tuñón, Iñaki, Moliner, Vicent and Allemann, Rudolf Konrad ORCID: 2013. Increased dynamic effects in a catalytically compromised variant of Escherichia coli dihydrofolate reductase. Journal of the American Chemical Society 135 (49) , pp. 18689-18696. 10.1021/ja410519h

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Isotopic substitution (15N, 13C, 2H) of a catalytically compromised variant of Escherichia coli dihydrofolate reduc-tase, EcDHFR-N23PP/S148A, has been used to investigate the effect of these mutations on catalysis. The reduction of the rate constant of the chemical step in the EcDHFR-N23PP/S148A catalysed reaction is essentially a consequence of an in-crease of the quasi-classical free energy barrier and to a minor extent of an increased number of recrossing trajectories on the transition state dividing surface. Since the variant enzyme is less well set up to catalyse the reaction a higher degree of active site reorganisation is needed to reach the TS. Although millisecond active site motions are lost in the variant, there is greater flexibility on the femtosecond timescale. The ‘dynamic knockout’ EcDHFR-N23PP/S148A is therefore a ‘dynamic knock-in’ at the level of the chemical step, and the increased dynamic coupling to the chemical coordinate is in fact detri-mental to catalysis. This finding is most likely applicable not just to hydrogen transfer in EcDHFR but also to other enzymatic systems.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Chemistry
Subjects: Q Science > QD Chemistry
Uncontrolled Keywords: kinetics, computational chemistry, biological chemistry, biophysics, dynamics
Additional Information: Online publication date: 19 November 2013.
Publisher: American Chemical Society
ISSN: 0002-7863
Funders: BBSRC
Date of First Compliant Deposit: 30 March 2016
Last Modified: 14 May 2023 16:56

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