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1e7y
From Proteopedia
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ACTIVE SITE MUTANT (D177->N) OF GLUCOSE 6-PHOSPHATE DEHYDROGENASE FROM LEUCONOSTOC MESENTEROIDES COMPLEXED WITH SUBSTRATE AND NADPH
Overview
The role of Asp-177 in the His-Asp catalytic dyad of glucose 6-phosphate, dehydrogenase from Leuconostoc mesenteroides has been investigated by a, structural and functional characterization of the D177N mutant enzyme. Its, three-dimensional structure has been determined by X-ray, cryocrystallography in the presence of NAD(+) and in the presence of, glucose 6-phosphate plus NADPH. The structure of a glucose 6-phosphate, complex of a mutant (Q365C) with normal enzyme activity has also been, determined and substrate binding compared. To understand the effect of, Asp-177 on the ionization properties of the catalytic base His-240, the pH, dependence of kinetic parameters has been determined for the D177N mutant, and compared to that of the wild-type enzyme. The structures give details, of glucose 6-phosphate binding and show that replacement of the Asp-177 of, the catalytic dyad with asparagine does not affect the overall structure, of glucose 6-phosphate dehydrogenase. Additionally, the evidence suggests, that the productive tautomer of His-240 in the D177N mutant enzyme is, stabilized by a hydrogen bond with Asn-177; hence, the mutation does not, affect tautomer stabilization. We conclude, therefore, that the absence of, a negatively charged aspartate at 177 accounts for the decrease in, catalytic activity at pH 7.8. Structural analysis suggests that the pH, dependence of the kinetic parameters of D177N glucose 6-phosphate, dehydrogenase results from an ionized water molecule replacing the missing, negative charge of the mutated Asp-177 at high pH. Glucose 6-phosphate, binding orders and orients His-178 in the D177N-glucose 6-phosphate-NADPH, ternary complex and appears to be necessary to form this water-binding, site.
About this Structure
1E7Y is a Single protein structure of sequence from Leuconostoc mesenteroides with , and as ligands. Active as Glucose-6-phosphate 1-dehydrogenase, with EC number 1.1.1.49 Known structural/functional Sites: , and . Full crystallographic information is available from OCA.
Reference
An examination of the role of asp-177 in the His-Asp catalytic dyad of Leuconostoc mesenteroides glucose 6-phosphate dehydrogenase: X-ray structure and pH dependence of kinetic parameters of the D177N mutant enzyme., Cosgrove MS, Gover S, Naylor CE, Vandeputte-Rutten L, Adams MJ, Levy HR, Biochemistry. 2000 Dec 12;39(49):15002-11. PMID:11106478
Page seeded by OCA on Sun Feb 3 09:38:04 2008
