Properties of Particulate and Detergent‐Solubilized Phospholipid N‐Methyltransferase Activity from Calf Brain

Publisher Website
Google Scholar
Abstract
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, > 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 were used as a basis for testing the hypothesis that the conversion of PME to PC is catalyzed by a single enzyme in calf brain. PME- and PDE-N-methyltransferase activities were similar, if not identical, with respect to the following: extractability with Triton X-100; pH optimum; response to divalent cations; apparent Km for S-adenosyl-L-methionine and Ki for S-adenosyl-L-homocysteine, sensitivity to N-ethylmaleimide and thermal inactivation at 55.degree. C. PME and PDE are methylated by the same enzyme or by 2 phospholipid N-methyltransferases having very similar properties.