Multiple Pathways for Triacylglycerol Biosynthesis in Streptomyces coelicolor

• Published in: Applied and Environmental Microbiology 2008-05, Vol.74 (9), p.2573-2582
• Main Authors: Arabolaza, Ana, Rodriguez, Eduardo, Altabe, Silvia, Alvarez, Hector, Gramajo, Hugo
• Format: Article
• Language: English
• Subjects: Acinetobacter; Acinetobacter - enzymology; Acyl Coenzyme A - metabolism; Acyltransferase; Acyltransferases - metabolism; Bacteria; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; Bacteriology; Biological and medical sciences; Cloning, Molecular; Enzymes; Fatty acids; Fundamental and applied biological sciences. Psychology; Gene Deletion; Gene Dosage; Gene Expression; Genes; Kinetics; Lipids; Metabolic Networks and Pathways; Microbiology; Physiology and Biotechnology; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Sequence Homology, Amino Acid; Soil microorganisms; Streptomyces coelicolor - enzymology; Streptomyces coelicolor - genetics; Streptomyces coelicolor - metabolism; Triglycerides - biosynthesis; Triglycerides - genetics
• ISSN: 0099-2240; 1098-5336; 1098-6596
• DOI: 10.1128/AEM.02638-07

The terminal reaction in triacylglyceride (TAG) biosynthesis is the esterification of diacylglycerol (DAG) with a fatty acid molecule. To study this reaction in Streptomyces coelicolor, we analyzed three candidate genes (sco0958, sco1280, and sco0123) whose products significantly resemble the recently identified wax ester synthase/acyl-coenzyme A (CoA):DAG acyltransferase (DGAT) from Acinetobacter baylyi. The deletion of either sco0123 or sco1280 resulted in no detectable decrease in TAG accumulation. In contrast, the deletion of sco0958 produced a dramatic reduction in neutral lipid production, whereas the overexpression of this gene yielded a significant increase in de novo TAG biosynthesis. In vitro activity assays showed that Sco0958 mediates the esterification of DAG using long-chain acyl-CoAs (C₁₄ to C₁₈) as acyl donors. The Km and Vmax values 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:DGAT (PDAT) activity in the wild type and in the triple mutant. This is the first time that a PDAT activity has been reported for bacteria, highlighting the extreme metabolic diversity of this industrially important soil microorganism.