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The effect of a Glu370Asp Mutation in Glutaryl-CoA Dehydrogenase on Proton Transfer to the Dienolate Intermediate
Structural highlights
DiseaseGCDH_HUMAN Defects in GCDH are the cause of glutaric aciduria type 1 (GA1) [MIM:231670. GA1 is an autosomal recessive metabolic disorder characterized by progressive dystonia and athetosis due to gliosis and neuronal loss in the basal ganglia.[1] [2] [3] [4] [5] [6] FunctionGCDH_HUMAN Catalyzes the oxidative decarboxylation of glutaryl-CoA to crotonyl-CoA and CO(2) in the degradative pathway of L-lysine, L-hydroxylysine, and L-tryptophan metabolism. It uses electron transfer flavoprotein as its electron acceptor. Isoform Short is inactive. Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedWe have determined steady-state rate constants and net rate constants for the chemical steps in the catalytic pathway catalyzed by the E370D mutant of glutaryl-CoA dehydrogenase and compared them with those of the wild-type dehydrogenase. We sought rationales for changes in these rate constants in the structure of the mutant cocrystallized with the alternate substrate, 4-nitrobutyric acid. Substitution of aspartate for E370, the catalytic base, results in a 24% decrease in the rate constant for proton abstraction at C-2 of 3-thiaglutaryl-CoA as the distance between C-2 of the ligand and the closest carboxyl oxygen at residue 370 increases from 2.9 A to 3.1 A. The net rate constant for flavin reduction due to hydride transfer from C-3 of the natural substrate, which includes proton abstraction at C-2, to N5 of the flavin decreases by 81% due to the mutation, although the distance increases only by 0.7 A. The intensities of charge-transfer bands associated with the enolate of 3-thiaglutaryl-CoA, the reductive half-reaction (reduced flavin with oxidized form of substrate), and the dienolate following decarboxylation are considerably diminished. Structural investigation suggests that the increased distance and the change in angle of the S-C1(=O)-C2 plane of the substrate with the isoalloxazine substantially alter rates of the reductive and oxidative half-reactions. This change in active site geometry also changes the position of protonation of the four carbon dienolate intermediate to produce kinetically favorable product, vinylacetyl-CoA, which is further isomerized to the thermodynamically stable normal product, crotonyl-CoA. The effect of a Glu370Asp mutation in glutaryl-CoA dehydrogenase on proton transfer to the dienolate intermediate.,Rao KS, Fu Z, Albro M, Narayanan B, Baddam S, Lee HJ, Kim JJ, Frerman FE Biochemistry. 2007 Dec 18;46(50):14468-77. Epub 2007 Nov 17. PMID:18020372[7] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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