Atomic details of near-transition state conformers for enzyme phosphoryl transfer revealed by MgF₃− rather than by phosphoranes

Authors

Nicola J. Baxter, University of Sheffield
Matthew W. Bowler, European Synchrotron Radiation Facility
Tooba Alizadeh, University of Sheffield
Matthew J. Cliff, University of Sheffield
Andrea M. Hounslow, University of Sheffield
Bin Wu, University of Nebraska - Lincoln
David B. Berkowitz, University of Nebraska - LincolnFollow
Nicholas H. Williams, University of Sheffield
G. Michael Blackburn, University of SheffieldFollow
Jonathan P. Waltho, University of SheffieldFollow

Date of this Version

3-9-2010

Citation

PNAS. March 9, 2010, vol. 107, no. 10, pp. 4555–4560.

Comments

Copyright 2010. Used by permission.

Abstract

Prior evidence supporting the direct observation of phosphorane intermediates in enzymatic phosphoryl transfer reactions was based on the interpretation of electron density corresponding to trigonal species bridging the donor and acceptor atoms. Close examination of the crystalline state of β-phosphoglucomutase, the archetypal phosphorane intermediate-containing enzyme, reveals that the trigonal species is not PO−3 , but is MgF−3 (trifluoromagnesate). Although MgF−3 complexes are transition state analogues rather than phosphoryl group transfer reaction intermediates, the presence of fluorine nuclei in near-transition state conformations offers new opportunities to explore the nature of the interactions, in particular the independent measures of local electrostatic and hydrogen-bonding distributions using 19F NMR. Measurements on three β-PGM-MgF−3 -sugar phosphate complexes show a remarkable relationship between NMR chemical shifts, primary isotope shifts, NOEs, cross hydrogen bond F⋯H-N scalar couplings, and the atomic positions determined from the highresolution crystal structure of the β-PGM-MgF₃- -G6P complex. The measurements provide independent validation of the structural and isoelectronic MgF₃− model of near-transition state conformations.