Phosphatidylethanolamine methyltransferase and phospholipid methyltransferase activities from Saccharomyces cerevisiae. Enzymological and kinetic properties

Abstract

In the yeast Saccharomyces cerevisiae, two membrane-associated enzymes catalyze the three-step methylation of Phosphatidylethanolamine (PE) to phosphatidylcholine (PC). Phosphatidylethanolamine methyltransferase (PEMT) catalyzes the first methylation reactions (PE → phosphatidylmonomethylethanolamine (PMME)) and phospholipid methyltransferase (PLMT) catalyzes the second two methylation reactions (PMME → phosphatidyldimethylethanolamine (PDME) → PC). Using gene disruption mutants of the S. cerevisiae OPI3 and CHO2 genes, we independently studied the enzymological properties of microsome-associated PEMT and PLMT, respectively. The enzymological properties of the enzymes differed with respect to their pH optima, cofactor requirements and thermal lability. For the PEMT reactions, the apparent K m values for PE and S-Adenosylmethionine (AdoMet) were 57 μM and 110 μM, respectively. For the PLMT reactions, the apparent K m values for PMME and PDME were 380 μM and 180 μM, respectively. The apparent K m values for AdoMet were 54 μM and 59 μM with PMME and PDME as substrates, respectively. S-Adenosylhomocysteine ( AdoHcy) was a competitive inhibitor of PEMT ( K i = 12 μM) and PLMT ( K) i = 57 μM and K i = 54 μM for PMME and PDME, respectively) with respect to AdoMet. AdoHcy was a noncompetitive inhibitor of PEMT ( K) i , = 160 μM) and PLMT K i = 320 μM and K i = 120 μM) with respect to PE and PMME and PDME, respectively.