Ligand geometry of the ternary complex of 5-enolpyruvylshikimate-3-phosphate synthase from rotational-echo double-resonance NMR

Biochemistry
L M McDowellJ Schaefer

Abstract

The 46-kDa enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase catalyzes the condensation of shikimate 3-phosphate (S3P) and phosphoenolpyruvate (PEP) to form EPSP. The reaction is inhibited by N-(phosphonomethyl)glycine (Glp), which, in the presence of S3P, binds to EPSP synthase to form a stable ternary complex. As part of a solid-state NMR characterization of this structure, we have used dipolar recovery at the magic angle (DRAMA) and rotational-echo double resonance (REDOR) to determine intra- and interligand internuclear distances. DRAMA was used to determine the single 31P-31P distance, while REDOR was used to determine one 31P-15N distance and five 31P-13C distances. These experimental distances were used as restraints in molecular dynamics simulations of an S3P-Glp complex to examine the geometry of the two ligands relative to one another in the ternary complex. The simulations were compared to unrestrained simulations of the EPSP synthase tetrahedral intermediate and its phosphonate analog. The results suggest that Glp is unlikely to bind in the same fashion as PEP, a conclusion that is consistent with recent studies that have questioned the role of Glp as a transition-state or intermediate analog.

References


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Citations

Aug 10, 2007·Journal of Biomolecular NMR·W Trent FranksChad M Rienstra
Nov 11, 1998·Solid State Nuclear Magnetic Resonance·J M GoetzJ Schaefer
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Mar 12, 2004·Annual Review of Plant Physiology and Plant Molecular Biology·Klaus M. Herrmann, Lisa M. Weaver
Feb 15, 2001·Proceedings of the National Academy of Sciences of the United States of America·E SchönbrunnW Kabsch
Aug 1, 2002·Proceedings of the National Academy of Sciences of the United States of America·Chad M RienstraRobert G Griffin
Nov 13, 1998·Proceedings of the National Academy of Sciences of the United States of America·T L BenzingerS C Meredith
Aug 15, 2000·Journal of Structural Biology·D G Lynn, S C Meredith
Aug 1, 1997·Journal of Magnetic Resonance·J M Goetz, J Schaefer
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Dec 5, 2006·Anais Da Academia Brasileira De Ciências·Luiz F L ReisMauro M Teixeira
Mar 17, 2004·Magnetic Resonance in Chemistry : MRC·Adriana Iuga, Eike Brunner

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