Abstract
Aldehyde dehydrogenases play crucial roles in the detoxification of exogenous and endogenous aldehydes by catalysing their oxidation to carboxylic acid counterparts. The present study reports characterization of two such isoenzymes from the yeast Saccharomyces cerevisiae var. boulardii (NCYC 3264), one mitochondrial (Ald4p) and one cytosolic (Ald6p). Both Ald4p and Ald6p were oligomeric in solution and demonstrated positive kinetic cooperativity towards aldehyde substrates. Wild-type Ald6p showed activity only with aliphatic aldehydes. Ald4p, on the contrary, showed activity with benzaldehyde along with a limited range of aliphatic aldehydes. Inspection of modelled structure of Ald6p revealed that a bulky amino acid residue (Met177, compared with the equivalent residue Leu196 in Ald4p) might cause steric hindrance of cyclic substrates. Therefore, we hypothesized that specificities of the two isoenzymes towards aldehyde substrates were partly driven by steric hindrance in the active site. A variant of wild-type Ald6p with the Met177 residue replaced by a valine was also characterized to address to the hypothesis. It showed an increased specificity range and a gain of activity towards cyclohexanecarboxaldehyde. It also demonstrated an increased thermal stability when compared with both the wild-types. These data suggest that steric bulk in the active site of yeast aldehyde dehydrogenases is partially responsible for controlling specificity.
Highlights
The multigene family of aldehyde dehydrogenases (EC 1.2.1.3) contributes primarily to acetaldehyde detoxification through its oxidation to acetate [1]
Mammalian aldehyde dehydrogenases are classified on the basis of kinetic mechanistic features into three classes of enzyme family: ALDH1, ALDH2 and ALDH3 being the representative members with rate-limiting steps in NADH dissociation, deacylation and hydride transfer respectively [11,12]
We investigated the specificity of two yeast aldehyde dehydrogenases for linear/branched-chain aliphatic aldehydes and cyclic/aromatic aldehydes as substrates
Summary
The multigene family of aldehyde dehydrogenases (EC 1.2.1.3) contributes primarily to acetaldehyde detoxification through its oxidation to acetate [1]. Mammalian aldehyde dehydrogenases are classified on the basis of kinetic mechanistic features into three classes of enzyme family: ALDH1, ALDH2 and ALDH3 being the representative members with rate-limiting steps in NADH dissociation, deacylation and hydride transfer respectively [11,12]. Whether intermediates or products, may be derived from the metabolism of drugs c 2017 The Author(s).
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