Phospholipid methylation in brain membrane preparations: kinetic mechanism.
The methylation reactions which convert phosphatidylethanolamine (PE) to phosphatidylcholine (PC) have been studied kinetically using exogenously added intermediates and crude membrane preparations from brain. The addition of exogenous PE resulted in no change in the methylation rates compared to that of endogenous PE. The addition of the two intermediates, monomethylphosphatidylethanolamine (PMME) and dimethylphosphatidylethanolamine (PDME), resulted in significantly increased rates of methylation and allowed the kinetic analysis of these latter two methylation reactions. The mechanism for this enzyme appears to be similar to human RBC (Reitz et al. (1989) J. Biol. Chem. 264, 8097-8106) which was a rapid-equilibrium random Bi-Bi sequential mechanism. There were some slight differences between the brain enzyme and that from the RBC, but there is little reason to suggest a fundamentally different mechanism. It is more likely that the differences may relate to an additional dead-end complex for the enzyme from brain such that saturation with AdoMet cannot eliminate AdoHcy inhibition. The KM values for the two phospholipid substrates were 41-44 microM and 39 microM for the methylation of PMME and PDME, respectively. The KM for S-adenosylmethionine (AdoMet) was 7-9 microM with PMME and 4 microM with PDME as the other substrates. The Ki(lipid) varied from 54 microM with PMME to 225 microM with PDME, and the Ki(AdoMet) was 11 microM with PMME and 21 microM with PDME. The product from the use of AdoMet, S-adenosylhomocysteine (AdoHcy), was shown to be a noncompetitive inhibitor of both lipid substrates as well as AdoMet. The methylation of PMME was somewhat higher in cerebellum and brain stem compared to cortex and striatum, but the methylation of PDME was similar in cerebellum, brain stem and cortex.
Reitz
R C
RC
Department of Biochemistry, School of Medicine, University of Nevada, Reno 89557.
Mead
D J
DJ
Welch
W H
WH
D000818:Animals; D001921:Brain / Q000201:enzymology* / Q000378:metabolism; D004912:Erythrocytes / Q000201:enzymology; D007700:Kinetics; D008297:Male; D008566:Membranes / Q000378:metabolism; D008745:Methylation; D008780:Methyltransferases / Q000378:metabolism*; D010714:Phosphatidylethanolamines / Q000378:metabolism / Q000494:pharmacology; D010743:Phospholipids / Q000378:metabolism*; D051381:Rats; D011916:Rats, Inbred F344; D012436:S-Adenosylmethionine / Q000378:metabolism
D010714:Phosphatidylethanolamines; D010743:Phospholipids; C032000:phosphatidyl-N-monomethylaminoethanol; C026508:phosphatidyldimethylethanolamine; D012436:S-Adenosylmethionine; D008780:Methyltransferases; C022352:phosphatidyl-N-monomethylethanolamine methyltransferase