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Probing active site geometry using high pressure and secondary isotope effects in an enzyme‐catalysed ‘deep’ H‐tunnelling reaction
Author(s) -
Hay Sam,
Pudney Christopher R.,
Sutcliffe Michael J.,
Scrutton Nigel S.
Publication year - 2010
Publication title -
journal of physical organic chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.325
H-Index - 66
eISSN - 1099-1395
pISSN - 0894-3230
DOI - 10.1002/poc.1653
Subject(s) - chemistry , quantum tunnelling , active site , kinetic isotope effect , high pressure , computational chemistry , stereochemistry , chemical physics , enzyme , thermodynamics , organic chemistry , atomic physics , deuterium , condensed matter physics , physics
We report the first study of the effects of hydrostatic pressure on α ‐2° KIEs for an enzyme‐catalysed H‐transfer reaction that occurs by ‘deep’ tunnelling. High pressure causes a significant decrease in the observed α ‐2° KIE on the pre‐steady‐state hydride transfer from NADH to FMN in the flavoprotein morphinone reductase. We have recently shown that high pressure causes a reduction in macroscopic reaction barrier width for this reaction. Using DFT vibrational analysis of a simple active site model, we posit that the decrease in α ‐2° KIE with pressure may arise due to a decrease in the vibrational coupling between the NADH primary (transferred) and secondary hydrogens in the ‘tunnelling ready configuration’, which more closely resembles the reactant state than the transition state. Copyright © 2010 John Wiley & Sons, Ltd.