In vivo and in vitro acylation of polypeptides in Vibrio harveyi: Identification of proteins involved in aldehyde production for bioluminescence

Incubation of soluble extracts from Vibrio harveyi with [³H]tetradecanoic acid (+ATP) resulted in the acylation of several polypeptides, including proteins with molecular masses near 20 kilodalton (kDa), and at least five polypeptides, in the 30- to 60-kDa range. However, in growing cells pulse-labeled in vivo with [³H]tetradecanoic acid, only three of these polypeptides, with apparent molecular masses of 54, 42 and 32-kDa, were specifically labeled. When extracts were acylated with [³H]tetradecanoyl coenzyme A, on the other hand, only the 32 kDa polypeptide was labeled. When luciferase-containing dark mutants of V. harveyi were investigated, acylated 32-kDa polypeptide was not detected in a fatty acid-stimulated mutant, whereas the 42-kDa polypeptide appeared to be lacking in a mutant defective in aldehyde synthesis. Acylation of both of these polypeptides also increased specifically during induction of bioluminescence in V. harveyi. These results suggest that the role of the 32-kDa polypeptide is to supply free fatty acids, whereas the 42-kDa protein may be responsible for activation of fatty acids for their subsequent reduction to form the aldehyde substrates of the bioluminescent reaction.