When radiolabeled serine is incubated with a particulate fraction from Saccharomyces cerevisiae, radioactivity is incorporated initially into phosphatidylserine and gradually appears in phosphatidylethanolamine. Because decarboxylation of phosphatidylserine is blocked by hydroxylamine, phosphatidylserine synthase can be assayed separately. The yeast phosphatidylserine synthase activity 1) exhibits a divalent cation requirement; 2) is stimulated by exogenous CDP-diolein (apparent Km = 0.17 mM); 3) has an apparent Km = 4 mM for L-serine; 4) has a neutral pH optimum; 5) is inhibited by p-hydroxymercuribenzoate; and 6) is reversible in the presence of 5'-CMP, but not 2'-CMP, 3'-CMP, or 5'-AMP. The phospholipid-synthesizing activity is solubilized with Triton X-100 and the enzymatic parameters have been compared with the particulate form of the enzyme. Detergent extracts catalyze the conversion of exogenous purified [31P]CDP-diglyceride to [32P]phosphatidylserine in the presence of Mn2+ and L-serine. Enzyme preparations from cells grown in the presence of choline, that have reduced phospholipid methylation activity (Waechter, C. J., Steiner, M. R., and Lester, R. L. (1969) J. Biol. Chem. 244, 3419-3422), also have substantially less phosphatidylserine synthase activity compared to identical preparations grown in the absence of choline. When choline, phosphocholine, CDP-choline, and phosphatidylcholine are present in vitro, there is no direct inhibitory effect on phosphatidylserine synthase activity. While the inclusion of choline in the growth medium caused a significant reduction in phosphatidylserine synthase activity, it did not appreciably effect the apparent Km values for L-serine and CDP-diglyceride. These results are consistent with choline-grown cells containing less phosphatidylserine synthase activity because of lower amounts of enzyme present or perhaps less active enzyme due to covalent modification.

• PMID: 6282872
• PubMed
• PubTator

Reference Type

Journal Article | Research Support, Non-U.S. Gov't

Authors

Carson MA, Atkinson KD, Waechter CJ

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