Detection of phospholipid biosynthetic enzyme activities in Saccharomyces cerevisiae by colony autoradiography.
The colony autoradiography method (C. R. H. Raetz (1975) Proc. Nat. Acad. Sci. USA 72, 2274-2278) was modified for the detection of CDP-diacylglycerol synthase, phosphatidylinositol synthase, phosphatidylserine synthase, and phosphatidylglycerophosphate synthase activities of Saccharomyces cerevisiae colonies on filter paper replica prints. Colonies were replica printed onto filter paper, permeabilized by air drying, and assayed for enzyme activities with labeled substrates. Autoradiograms of replica prints, following enzyme assays, showed dark halos indicating the enzymatic synthesis of labeled phospholipid products. The method was also used to detect a cho 1 mutant defective in phosphatidylserine synthase and a strain that overproduces phosphatidylserine synthase. The method should become a valuable tool in isolating yeast strains defective in phospholipid biosynthetic enzyme activities and strains with overproduced enzyme activities.
D001345:Autoradiography; D004798:Enzymes / Q000235:genetics; D009154:Mutation; D010743:Phospholipids / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics
D004798:Enzymes; D010743:Phospholipids