CATALYTIC SELF-ACYLATION OF TYPE-II POLYKETIDE SYNTHASE ACYL CARRIER PROTEINS

Background: Aromatic polyketides are synthesised in streptomycetes by the successive condensation of simple carboxylic acids, catalysed by m ultienzyme complexes - the polyketide synthases (PKSs). Polyketide ass embly intermediates are covalently linked as thioesters to the holo-ac yl carrier protein (ACP) subunit of these type II PKSs. The ACP is pri med for chain elongation by the transfer of malonate from malonyl CoA. Malonylation of fatty acid synthase (FAS) ACPs is catalysed by specif ic malonyl transferase (MT) enzymes. The type II PKS gene clusters app arently lack genes encoding such MT proteins, however. It has been pro posed that the MT subunit of the FAS in streptomycetes catalyses malon ylation of both FAS and PKS ACPs in vivo. Results: We demonstrate that type II PKS ACPs catalyse self-malonylation upon incubation with malo nyl CoA in vitro. The self-malonylation reaction of the actinorhodin C 17S holo-ACP has a K-m for malonyl CoA of 219 mu M and a k(cat) of 0.3 4 min(-1). Complete acylation of the PKS ACPs was observed with malony l, methylmalonyl and acetoacetyl CoAs. No reaction was observed with a cetyl and butyryl CoAs and FAS ACPs did not react with any of the subs trates. Recombinant FAS MT from Streptomyces coelicolor did not accele rate the rate of malonylation. Conclusions: The catalytic self-acylati on of type II PKS ACPs is an unprecedented reaction, We propose a reac tion mechanism in which conserved arginines form a salt bridge with th e acyl moiety and sequester it from bulk solvent. This work suggests t hat the beta-ketoacyl synthase, chain length factor and ACP may consti tute a truly minimal PKS in vivo.