Abstract
Methionine synthase is a key enzyme in the methionine cycle that catalyzes the transmethylation of homocysteine to methionine in a cobalamin-dependent reaction that utilizes methyltetrahydrofolate as a methyl group donor. Cob(I)alamin, a supernucleophilic form of the cofactor, is an intermediate in this reaction, and its reactivity renders the enzyme susceptible to oxidative inactivation. In bacteria, an NADPH-dependent two-protein system comprising flavodoxin reductase and flavodoxin, transfers electrons during reactivation of methionine synthase. Until recently, the physiological reducing system in mammals was unknown. Identification of mutations in the gene encoding a putative methionine synthase reductase in the cblE class of patients with an isolated functional deficiency of methionine synthase suggested a role for this protein in activation (Leclerc, D., Wilson, A., Dumas, R., Gafuik, C., Song, D., Watkins, D., Heng, H. H. Q., Rommens, J. M., Scherer, S. W., Rosenblatt, D. S., and Gravel, R. A. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 3059-3064). In this study, we have cloned and expressed the cDNA encoding human methionine synthase reductase and demonstrate that it is sufficient for supporting NADPH-dependent activity of methionine synthase at a level that is comparable with that seen in the in vitro assay that utilizes artificial reductants. Methionine synthase reductase is a soluble, monomeric protein with a molecular mass of 78 kDa. It is a member of the family of dual flavoproteins and is isolated with an equimolar concentration of FAD and FMN. Reduction by NADPH results in the formation of an air stable semiquinone similar to that observed with cytochrome P-450 reductase. Methionine synthase reductase reduces cytochrome c in an NADPH-dependent reaction at a rate (0.44 micromol min(-1) mg(-1) at 25 degrees C) that is comparable with that reported for NR1, a soluble dual flavoprotein of unknown function, but is approximately 100-fold slower than that of P-450 reductase. The K(m) for NADPH is 2.6 +/- 0.5 microm, and the K(act) for methionine synthase reductase is 80.7 +/- 13.7 nm for NADPH-dependent activity of methionine synthase.
Highlights
Homocysteine is a key junction metabolite in the methionine cycle that is formed by the hydrolysis of S-adenosylhomocysteine, the product of S-adenosylmethionine-dependent methyl
Methionine synthase is a key enzyme in the methionine cycle that catalyzes the transmethylation of homocysteine to methionine in a cobalamin-dependent reaction that utilizes methyltetrahydrofolate as a methyl group donor
The mammalian methionine synthase is a methylcobalamindependent enzyme that catalyzes the successive transfer of a methyl group from CH3-H4folate1 to the cob(I)alamin form of the cofactor and from methylcobalamin to homocysteine to form methionine and tetrahydrofolate as products (Fig. 1) [6]
Summary
Homocysteine is a key junction metabolite in the methionine cycle that is formed by the hydrolysis of S-adenosylhomocysteine, the product of S-adenosylmethionine-dependent methyl-. The transmethylation reactions catalyzed by methionine synthase or betaine homocysteine methyltransferase salvage homocysteine back to the methionine cycle. The mammalian methionine synthase is a methylcobalamindependent enzyme that catalyzes the successive transfer of a methyl group from CH3-H4folate to the cob(I)alamin form of the cofactor and from methylcobalamin to homocysteine to form methionine and tetrahydrofolate as products (Fig. 1) [6]. The reactivity of the intermediate cob(I)alamin form of the enzyme renders the enzyme susceptible to oxidation, resulting in adventitious formation of cob(II)alamin The latter represents an inactive form of the enzyme that can be returned to the catalytic cycle by a reductive methylation reaction that utilizes S-adenosylmethionine as a methyl group donor. Two studies, using genetic and biochemical approaches, respectively, identified two pathways for mammalian methionine synthase activation (Fig. 2). Biochemical characterization of the gene product and its ability to activate methionine synthase were not investigated
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
