Catalytic properties of a bacterial acylating acetaldehyde dehydrogenase: evidence for several active oligomeric states and coenzyme A activation upon binding.

摘要: Until the last decade, two unrelated aldehyde dehydrogenase (ALDH) superfamilies, i.e. phosphorylating and non-phosphorylating were known to catalyze oxidation of aldehydes activated or non-activated acids. However, a third one was discovered by crystal structure bifunctional enzyme 4-hydroxy-2-ketovalerate aldolase/acylating acetaldehyde (DmpFG) from Pseudomonas sp. strain CF600 (Manjasetty et al., Proc. Natl. Acad. Sci. USA 100 (2003) 6992-6997). Indeed, DmpF exhibits CoA-dependent ALDH activity, but is structurally related superfamily. In this study, we undertook characterization catalytic structural properties MhpEF Escherichia coli, an ortholog DmpFG in which MhpF converts acetaldehyde, produced cleavage MhpE, into acetyl-CoA. The kinetic data obtained under steady-state pre-steady-state conditions show that dehydrogenase, MhpF, active as monomer, unique feature relative superfamilies. Our results also reveal are not dependent on its oligomeric state, supporting hypothesis catalytically independent entity. Moreover, transthioesterification shown be rate-limiting and, when compared with chemical model, efficiency increased 10(4)-fold. Therefore, CoA binding increases reactivity optimizes positioning thioacylenzyme intermediate, thus enabling formation efficient deacylation complex.