CHARACTERIZATION OF THE yhdO GENE OF Bacillus subtilis, THAT CODES FOR A PUTATIVE ACYLTRANSFERASE

PAOLETTI L, SCHUJMAN GE, DE MENDOZA D

Villa Carlos Paz, Córdoba

Congreso; XXXVIIIReunión Anual de la Solciedad Argentina de Bioquímica y Biología Molecular (SAIB); 2002

Solciedad Argentina de Bioquímica y Biología Molecular

The first step in phospholipid biosynthesis is the transfer of the acyl moiety of two acyl-ACPs to glycerol-3-P to produce phosphatidic acid, catalyzed by enzymes called acyltransferases. In order to characterize this process in a Gram-positive bacterium, we analyzed the genome of Bacillus. subtilis and identified only one gene with homology to known acyl-ACP transacylases. To study the function of this gene, called yhdO, we obtained the strain LP15 in which the chromosomal copy of the gene was under the Pspac promoter, whose expression is induced in the presence of IPTG. Cultures of this strain were unable to grow in the absence of the inductor, indicating that the yhdO gene is essential in B. subtilis. Cultures of strain PL15 were labeled with [2-14C]-acetate in the presence or absence of IPTG, and lipids were extracted and identified. We found in this study that the absence of the inductor caused the accumulation of  large amounts of free fatty acids and that phospholipds synthesis was blocked thus, suggesting that yhdO gene is involved in the transacylation of fatty acids to glycerol-3-P. We studied the promoter region of yhdO gene by primer extension, gel shift assays and lacZ transcriptional fusions and determined that the expression of this gene is coordinately regulated with fatty acid biosynthesis by the FapR transcriptional repressor. This is the first characterization of a gene that codes for an acyl-ACP transacylase from a Gram positive bacterium.