The studies outlined in the present proposal are designed to provide information on the cellular regulatory mechanisms employed by phototrophically growing cells of Rhodopseudomonas sphaeroides to coordinate phospholipid synthesis with cell growth and membrane assembly. Studies on the purification of the sn-glycerol-3-phosphate acyltransferase from phototrophically grown cells will be conducted as a prerequisite to developing methods for monitoring the kinetics of synthesis of this enzyme in synchronously dividing cell populations. Studies on the identification and cloning of the structural gene for the R. sphaeroides sn-glycerol-3-phosphate acyltransferase will also be conducted. In vivo studies conducted with synchronously dividing cell populations, and cell populations subjected to immediate high-to-low light transitions, will be utilized to identify the level at which the temporal control and light-mediated control of phospholipid synthesis are exerted, respectively. By comparing the patterns of in vivo rates of cellular fatty acid and phospholipid synthesis, and by monitoring the quantitative and qualitative changes in the in vivo levels of acyl carrier protein (ACP) and acyl-ACP, regulatory mechanisms operative at the level of fatty acid synthesis will be distinguished from control mechanisms which function at the level of phospholipid synthesis. As warranted, these studies will be accompanied by direct, in vitro studies of enzymes involved in fatty acid and/or phospholipid biosynthesis in order to specifically identify the site and nature of the observed regulation.
