Abstract
The membrane localization and properties of the Rhodopseudomonas sphaeroides sn-glycerol-3-phosphate acyl transferase have been examined utilizing enzymatically prepared acyl-acyl carrier protein (acyl-ACP) substrates as acyl donors for sn-glycerol-3-phosphate acylation. Studies conducted with membranes prepared from chemotrophically and phototrophically grown cells show that sn-glycerol-3-phosphate acyl transferase activity is predominantly (>80%) associated with the cell's cytoplasmic membrane. Enzyme activity associated with the intracytoplasmic membranes present in phototrophically grown R. sphaeroides was within the range attributable to cytoplasmic membrane contamination of this membrane fraction. Enzyme activity was optimal at 40°C and pH 7.0 to 7.5 and required the presence of magnesium. No enzyme activity was observed with any of the long-chain acyl-CoA substrates examined. Vaccenoyl-ACP was the preferred acyl-ACP substrate and both vaccenoyl-ACP and palmitoyl-ACP were independently utilized to produce lysophosphatidic and phosphatidic acids. With either vaccenoyl-ACP or palmitoyl-ACP as sole acyl donor substrate, the lysophosphatidic acid formed was primarily 1-acylglycerol-3 -phosphate and the apparent Km for sn-glycerol-3-phosphate utilization was 100 μM. In addition, the acyl carrier protein (ACP) has been purified from photoheterotrophically grown cells of R. sphaeroides. The ACP preparation was determined to be >95% homogeneous as judged by native and sodium dodecylsulfate gel electrophoresis and by N-terminal amino acid analysis. The results of amino acid compositional analysis revealed that the ACP contains approximately 75 amino acids, lacks the amino acids tyrosine and tryptophan and has a calculated minimum molecular weight of 8,699. In contrast to E. coli ACP, R. sphaeroides ACP apparently binds sodium dodecylsulfate normally and displays an Mr of 8,700 when analyzed by gel electrophoresis. The protein contains equimolar quantities of β-alanine and taurine, indicating the presence of the characteristic 4'-phosphopantetheine prosthetic group, has an experimentally determined pi of 3.8 and a calculated partial specific volume of 0.732 ml/g...
Cooper, Cynthia LaFaye (1985). Localization and characterization of the sn-glycerol-3-phosphate acyltransferase and purification of the acyl carrier protein from Rhodopseudomonas sphaeroides. Texas A&M University. Texas A&M University. Libraries. Available electronically from
https : / /hdl .handle .net /1969 .1 /DISSERTATIONS -447647.