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Methionine synthase reductase undergoes phosphate‐sensitive loss of flavin mononucleotide upon dilution in vitro
Author(s) -
Elmore Calvin Lee
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.634.2
Subject(s) - flavin mononucleotide , methionine synthase , chemistry , thymidylate synthase , flavoprotein , flavin group , biochemistry , cofactor , methionine , flavin adenine dinucleotide , enzyme , reductase , dihydrofolate reductase , transmethylation , biosynthesis , stereochemistry , biology , amino acid , fluorouracil , chemotherapy , genetics
Hyperhomocyst(e)inemia is a metabolic derangement that is linked to the distribution of folate pools, which provide one‐carbon units for biosynthesis of purines and thymidylate and for remethylation of homocysteine to form methionine. In humans, methionine synthase deficiency results in the accumulation of methyltetrahydrofolate at the expense of folate derivatives required for purine and thymidylate biosynthesis. Methionine synthase reductase (MSR) is absolutely required for methionine synthase activity in vivo. MSR is a dual flavoprotein, containing one mol of FAD and one mol of FMN per mol of enzyme. While free FMN is highly fluorescent, enzyme‐bound FMN fluorescence is quenched. Flavin mononucleotide (FMN) binding in MSR is weak compared to other dual flavin‐containing enzymes (K d = ~85 μM; 0 mM P i ), especially in the presence of phosphate (K d = ~450 μM; 50 mM P i ) at constant ionic strength. Release of FMN from MSR is associated with loss of enzyme activity, as determined by cytochrome c reductase assay, while FMN‐reconstituted enzyme regains all activity. We found that the presence of 50 mM phosphate increased the overall loss of reductase activity upon dilution by ~50% and decreased the rate of rebinding of FMN ~5‐fold compared to phosphate free buffer. These findings point to a mechanism by which MSR may be inactivated under conditions that limit the availability of FMN in vivo in a phosphate sensitive manner.