Properties of particulate and detergent-solubilized phospholipid N-methyltransferase activity from calf brain.
Calf brain membranes catalyze the enzymatic transfer of [CH3-3H]methyl groups from S-adenosyl-L-[CH3-3H]methionine into endogenous phosphatidyl-N-methylethanolamine (PME), phosphatidyl-N,N-dimethylethanolamine (PDE), and phosphatidylcholine (PC). Phospholipid N-methylation can be stimulated by the addition of exogenous PME or PDE, added in aqueous dispersions with sodium taurocholate. When membranes are incubated in the presence of exogenous PME, [CH3-3H]PDE represents 86% of the labeled phospholipid products. When exogenous PME is replaced by PDE, 91% of the label is incorporated into PC. Thus, under these in vitro conditions it is possible to assay PME- and PDE-N-methyltransferase activity separately. The calf brain phospholipid N-methyltransferase activity has also been solubilized by treating the membranes ultrasonically in the presence of Triton X-100 and 10 mM monothioglycerol. When the detergent extracts are incubated in the presence of exogenous PME, [CH3-3H]PDE represents 86% of the enzymatically labeled products. In the presence of exogenous PDE, more than 97% of the label is incorporated into PC. Optimal conditions for the membrane-bound and detergent-solubilized PME- and PDE-N-methyltransferase activity have been established. These conditions have been used as a basis for testing the hypothesis that the conversion of PME to PC is catalyzed by a single enzyme in calf brain. In these studies, PME- and PDE-N-methyltransferase activities have been found to be similar, if not identical, with respect to: (1) extractability with Triton X-100; (2) pH optimum; (3) response to divalent cations; (4) apparent Km for S-adenosyl-L-methionine and K1 for S-adenosyl-L-homocysteine, (5) sensitivity to N-ethylmaleimide; and (6) thermal inactivation at 55 degrees. Overall, these results are consistent with the conclusion that in calf brain, PME and PDE are methylated by the same enzyme or by two phospholipid N-methyltransferases having very similar properties.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D007700:Kinetics; D008780:Methyltransferases / Q000378:metabolism*; D010713:Phosphatidylcholines / Q000378:metabolism; D050918:Phosphatidylethanolamine N-Methyltransferase; D010714:Phosphatidylethanolamines / Q000378:metabolism*; D012436:S-Adenosylmethionine / Q000378:metabolism; D012995:Solubility; D013656:Taurocholic Acid
D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; C032000:phosphatidyl-N-monomethylaminoethanol; C026508:phosphatidyldimethylethanolamine; D013656:Taurocholic Acid; D012436:S-Adenosylmethionine; D008780:Methyltransferases; D050918:Phosphatidylethanolamine N-Methyltransferase