Caractérisation enzymatique de la Méthylmalonate semialdéhyde déshydrogénase de Bacillus subtilis

Abstract : Homotetrameric methylmalonate semialdehyde dehydrogenase (MSDH) from Bacillus subtilis catalyzes the NAD+-dependent oxidation of methylmalonate semialdehyde into propionyl-Coenzyme A via a two-step mechanism. A detailed mechanistic characterization of the MSDH-catalyzed reaction has been carried out. The results show that NAD binding elicits a structural imprinting of the apo-enzyme. The enzyme exhibits a half-of-the-site reactivity with two subunits active per tetramer. The results support also a sequential C302 activation process with a pKapp shift from ~ 8.8 in the apo-form to 8.0 in the binary complex and finally ~ 5.5 in the ternary complex. The rate-limiting step is shown to be associated with the beta-decarboxylation process which occurs on the thioacylenzyme intermediate after NADH release and before the transthioesterification step. Binding of the substrate is favoured by stabilizing interactions between its carboxylate group and the invariant residues R124 and R301.
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Claire Chaumeil-Stines. Caractérisation enzymatique de la Méthylmalonate semialdéhyde déshydrogénase de Bacillus subtilis. Sciences agricoles. Université Henri Poincaré - Nancy 1, 2005. Français. ⟨NNT : 2005NAN10186⟩. ⟨tel-01748077⟩

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