Involvement of Acyl Carrier Protein in Acylation of Glycerol 3-Phosphate in Clostridium butyricum: II. EVIDENCE FOR THE PARTICIPATION OF ACYL THIOESTERS OF ACYL CARRIER PROTEIN

Abstract

Abstract 1. In order to examine the involvement of acyl carrier protein in the acylation of glycerophosphate by bacterial extracts, a series of experiments with radioactive glycerol 3-phosphate, potential acyl donors, and enzyme preparations from Clostridium butyricum were carried out. 2. The conversion of 14C-glycerophosphate to lipid, mainly lysophosphatidic acid, in the presence of a particulate fraction from C. butyricum was found to be dependent on the addition of acyl carrier protein (ACP) isolated from either C. butyricum or Escherichia coli. The apparent Km was 1.2 x 10-6 m for C. butyricum ACP and 3.4 x 10-6 m for E. coli ACP. Since no source of acyl groups was added in this series of experiments, it is postulated that the acyl groups transferred to glycerophosphate were derived from the particles. 3. With this particulate fraction, chemically synthesized 3H-palmityl-ACP also markedly stimulated the acylation of glycerol 3-phosphate and was an efficient donor of palmitate in the synthesis of lysophosphatidic acid. Palmityl coenzyme A had little effect on glycerophosphate acylation with the particulate fraction as the sole enzyme source. 4. A two-fold stimulation of glycerophosphate acylation by palmityl-CoA was observed when a soluble protein fraction from C. butyricum was added to the particulate fraction. Under these conditions some transfer of labeled fatty acids from 3H-acyl-CoA to 14C-glycerophosphate was observed. The lysophosphatidic acid isolated had 0.3 3H-acyl group per 14C-glycerophosphate. Presumably, the remainder of the acyl groups was derived from the enzyme sources. When ACP was added, the labeled fatty acids derived from acyl-CoA were further diluted by endogenous fatty acids. 5. Sodium lauryl sulfate also stimulated glycerophosphate acylation in the presence of particles from C. butyricum plus the soluble protein fraction. Both palmityl-CoA and sodium lauryl sulfate stimulated glycerophosphate acylation more at 25° than at 30°. These results suggest that some of the effects of palmityl-CoA in this system are the result of its surfactant properties.