Multiple Pathways for Triacylglycerol Biosynthesis in Streptomyces coelicolor

Microbiology Division, IBR (Instituto de Biología Molecular y Celular de Rosario), Consejo Nacional de Investigaciones Científicas y Técnicas, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, (S2002LRK) Rosario, Argentina; Centro Regional de Investigación y Desarrollo Científico Tecnológico (CRIDECIT) Facultad de Ciencias Naturales Universidad Nacional de la Patagonia San Juan Bosco Km 4- Ciudad Universitaria (9000) Comodoro Rivadavia (Chubut), Argentina

^* To whom correspondence should be addressed. Email: gramajo{at}ibr.gov.ar.

	Abstract

The terminal reaction in triacylglyceride (TAG) biosynthesis is esterification of diacylglycerol (DAG) with a fatty acid molecule. To study it in Streptomyces coelicolor we analyzed three candidate genes (sco0958, sco1280, sco0123) whose products significantly resemble the recently identified wax ester synthase/acyl-CoA:diacylglycerol acyltransferase (WS/DGAT) from Acinetobacter baylyi. Deletion of either sco0123 or sco1280 resulted in no detectable decrease in TAG accumulation. In contrast, deletion of sco0958 produced a dramatic reduction in neutral lipid production, whereas over-expression of this gene yielded a significant increase in de novo TAG biosynthesis. In vitro activity assays showed that Sco0958 mediates esterification of DAG using long-chain acyl-CoAs (C₁₄ to C₁₈) as acyl donors. The K_m and V_max of this enzyme for myristoyl-CoA were 45 µM and 822 nmol mg^-1 min^-1, respectively. Significantly, the triple mutant strain was not completely devoid of storage lipids, indicating the existence of alternative TAG biosynthetic routes. We present strong evidence demonstrating that the residual production of TAG in this mutant strain is mediated, at least in part, by an acyl CoA-dependent pathway, since the triple mutant still exhibited DGAT activity. More importantly, there was substantial phospholipid: diacylglycerol acyltransferase activity (PDAT) in the wild type and in the triple mutant. This is the first time that a PDAT activity has been reported in bacteria, highlighting the extreme metabolic diversity of this industrially important soil microorganism.