J Mol Biol. 2001 Jun 22;309(5):1209-18.

[DOI Link]

Structural assembly of the active site in an aldo-keto reductase by NADPH cofactor.

Institute of Molecular Biophysics and Department of Chemistry, Florida State University, Tallahassee, FL 32306-4380, USA.

A 1.9 A resolution X-ray structure of the apo-form of Corynebacterium 2,5-diketo-d-gluconic acid reductase A (2,5-DKGR A), a member of the aldo-keto reductase superfamily, has been determined by molecular replacement using the NADPH-bound form of the same enzyme as the search model. 2,5-DKGR A catalyzes the NADPH-dependent stereo-specific reduction of 2,5-diketo-d-gluconate (2,5-DKG) to 2-keto-l-gulonate, a precursor in the industrial production of vitamin C. An atomic-resolution structure for the apo-form of the enzyme, in conjunction with our previously reported high-resolution X-ray structure for the holo-enzyme and holo/substrate model, allows a comparative analysis of structural changes that accompany cofactor binding. The results show that regions of the active site undergo coordinated conformational changes of up to 8 A. These conformational changes result in the organization and structural rearrangement of residues associated with substrate binding and catalysis. Thus, NADPH functions not only to provide a hydride ion for catalytic reduction, but is also a critical structural component for formation of a catalytically competent form of DKGR A. Copyright 2001 Academic Press.

PMID: 11399090 [PubMed - indexed for MEDLINE]

This publication is one of the several that describes a structure solved either at the Kasha Laboratory, Institute of Molecular Biophysics or in collaboration with the Institute Faculty. The data used for this structure determination came in full or part from the Macromolecular X-Ray Crystallography Facility.