Multiple pathways for triacylglycerol biosynthesis in Streptomyces coelicolor.
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 (C(14) to C(18)) as acyl donors. The K(m) and V(max) values of this enzyme for myristoyl-CoA were 45 muM 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.
Arabolaza
Ana
A
Microbiology Division, 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, Rosario, Argentina.
Rodriguez
Eduardo
E
Altabe
Silvia
S
Alvarez
Hector
H
Gramajo
Hugo
H
D000150:Acinetobacter / Q000201:enzymology; D000214:Acyl Coenzyme A / Q000378:metabolism; D000217:Acyltransferases / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D003001:Cloning, Molecular; D017353:Gene Deletion; D018628:Gene Dosage; D015870:Gene Expression; D007700:Kinetics; D053858:Metabolic Networks and Pathways; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D048372:Streptomyces coelicolor / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D014280:Triglycerides / Q000096:biosynthesis* / Q000235:genetics
D000214:Acyl Coenzyme A; D001426:Bacterial Proteins; D011994:Recombinant Proteins; D014280:Triglycerides; C040349:S-tetradecanoyl-coenzyme A; D000217:Acyltransferases; C426836:phospholipid diacylglycerol acyltransferase