Study of the enzymes involved in the first steps of the phospholipid biosynthesis in Bacillus subtilis

PAOLETTI L, SCHUJMAN GE, DE MENDOZA D

Rosario- Santa Fe

Congreso; XLII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular- SAIB; 2006

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

In E. coli two acyltransferases, PlsB and PlsC, are required for phosphatidic acid production. A different pathway has been described in Streptococcus pneumoniae. The pathway involves two enzymes, encoded by plsX and plsY genes, which catalyze the reaction of acylation of G3P using a new intermediate, fatty acyl-phosphate. In B. subtilis there are genes with homology to plsX, plsY and plsC, but no plsB homolog could be identified. To determine the functions of the enzymes encoded by plsXb, plsYb and plsCb in B. subtilis, conditional mutants were constructed. The analysis of the mutants indicated that the expression of each gene is essential for the growth of B. subtilis. The analysis of the lipids extracted from cultures of the conditional mutants without inducer indicated that phospholipids biosynthesis was blocked in the three mutants, and fatty acids accumulated in the plsYb and plsCb conditional mutants. The subcellular localization of PlsXb and PlsCb was analyzed by immunofluorescence microscopy (IFM). In these experiments PlsCb and PlsXb were found homogeneously distributed in the cytoplasmic membrane, but PlsXb is a soluble protein as determinated by western blot. These results suggest that B. subtilis uses the PlsX/Y pathway for the phospholipids biosynthesis, and that a putative protein attaches in vivo PlsXb to the cell membrane.