Methionine synthase inactivation by nitrous oxide during methionine loading of normal human fibroblasts. Homocysteine remethylation as determinant of enzyme inactivation and homocysteine export.
Nitrous oxide inactivates the enzyme methionine synthase by oxidation of enzyme bound cobalamin, which is formed from the cofactor methylcobalamin during the catalytic cycle. The resulting inhibition of homocysteine remethylation increases the homocysteine efflux and thereby the level of extracellular homocysteine, both in patients and cultured cells. In the present work we measured the kinetics of enzyme inactivation and homocysteine export rate in two human fibroblast cell lines exposed to nitrous oxide and cultured in the presence of low to supraphysiological concentrations (15-100 microM) of methionine. Both the rate and extent of methionine synthase inactivation were reduced by increasing methionine concentration in the culture medium. In cells not exposed to nitrous oxide, methionine increased the homocysteine export rate in a dose-dependent manner. Nitrous oxide increased the export at low methionine concentrations, so that for treated cells the export was high and essentially independent of the extracellular methionine level. Neither methionine nor nitrous oxide significantly affected the amount of S-adenosylmethionine or folate in these cells. These data agree with methionine synthase as a low Km and methionine conserving enzyme, highlight the importance of methionine synthase activity as a determinant of homocysteine export and point to the possibility of protecting the enzyme by reducing catalytic turnover through product inhibition.
Christensen
B
B
Department of Pharmacology and Toxicology, University of Bergen, Norway.
Ueland
P M
PM
D013766:5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase / Q000037:antagonists & inhibitors*; D002455:Cell Division / Q000502:physiology; D002460:Cell Line; D004789:Enzyme Activation; D005110:Extracellular Space / Q000378:metabolism; D005333:Fetus / Q000166:cytology; D005347:Fibroblasts / Q000187:drug effects* / Q000201:enzymology* / Q000378:metabolism; D005492:Folic Acid / Q000378:metabolism; D006710:Homocysteine / Q000378:metabolism / Q000493:pharmacokinetics*; D006801:Humans; D007223:Infant; D008168:Lung / Q000196:embryology; D008297:Male; D008715:Methionine / Q000378:metabolism* / Q000494:pharmacology*; D008745:Methylation / Q000187:drug effects; D009609:Nitrous Oxide / Q000494:pharmacology*; D012436:S-Adenosylmethionine / Q000378:metabolism
D006710:Homocysteine; D012436:S-Adenosylmethionine; D005492:Folic Acid; D008715:Methionine; D013766:5-Methyltetrahydrofolate-Homocysteine S-Methyltransferase; D009609:Nitrous Oxide