Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: KEGG:D02011 (FAD)
5,530 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the presence of 5-methoxytryptamine (5-MeOT), 5-methyltetrahydrofolic acid (5-MTHF) yields 6-methoxy-1,2,3,4-tetrahydro-beta-carboline (6-MeOTHbetaC) in rat brain extracts, possibly via formaldehyde formation catalyzed by methylenetetrahydrofolate reductase. The formation of 6-MeOTHbetaC in selected brain regions, ranging from 452 +/- 40 pmol formed per mg protein per hour in corpus striatum to 119 +/- 17 pmol in cingulate cortex, is significantly correlated with the regional distribution of 1,2,3,4-tetrahydro-beta-carboline (THbetaC) formed from 5-MTHF and tryptamine (r = 0.76, p less than 0.01) as well as that of methylene-beta-phenylethylimine (MbetaphiEI) from 5-MTHF and beta-phenylethylamine (betaphiEA; r = 0.90, p less than 0.01). FAD enhances the activity, lowering both Vmax and Km values with respect to 5-MeOT and Vmax, but not Km, with respect to 5-MTHF.
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PMID:Regional formation of 6-methoxy-1,2,3,4-tetrahydro-beta-carboline in rat brain extract. 119 18

The methylenetetrahydrofolate reductase from the carbon-monoxide-utilizing homoacetogen Peptostreptococcus productus (strain Marburg) has been purified to apparent homogeneity. The purified enzyme catalyzed the oxidation of NADH with methylenetetrahydrofolate as the electron acceptor at a specific activity of 380 mumols.min-1 mg protein-1 (37 degrees C; pH 5.5). The apparent Km for NADH was near 10 microM. The apparent molecular mass of the enzyme was determined by gel filtration to be approximately 250.0 kDa. The enzyme consists of eight identical subunits with a molecular mass of 32 kDa. It contains 4 FAD/mol octamer which were reduced by the enzyme with NADH as the electron donor; iron could not be detected. Oxygen had no effect on the enzyme. Ultracentrifugation of cell extracts revealed that about 40% of the enzyme activity was recovered in the particulate fraction, suggesting that the enzyme is associated with the membrane. The enzyme also catalyzed the methylenetetrahydrofolate reduction with methylene blue as an artificial electron donor. The oxidation of methyltetrahydrofolate was mediated with methylene blue as the electron acceptor; neither NAD+ nor viologen dyes could replace methylene blue in this reaction. NADP(H) or FAD(H2) were not used to substrates for the reaction in either direction. The activity of the purified enzyme, which was proposed to be involved in sodium translocation across the cytoplasmic membrane, was not affected by the absence or presence of added sodium. The properties of the enzyme differ from those of the ferredoxin-dependent methylenetetrahydrofolate reductase of the homoacetogen Clostridium formicoaceticum and of the NADP(+)-dependent reductase of eucaryotes investigated so far.
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PMID:Purification and properties of a NADH-dependent 5,10-methylenetetrahydrofolate reductase from Peptostreptococcus productus. 220 95

Methylenetetrahydrofolate reductase from human cadaver liver was purified to homogeneity. The purified enzyme had a molecular mass of 150 kDa. On SDS-polyacrylamide gel electrophoresis it was dissociated into a single fragment with a molecular mass of 39 kDa. In contrast, fresh lymphocyte enzyme extract showed a major band with a molecular mass of 75 kDa and a minor band of 39 kDa. Fresh liver enzyme was inhibited by S-adenosylmethionine while the purified enzyme from human cadaver liver was not inhibited. These observations suggest that human methylenetetrahydrofolate reductase is composed of two identical subunits of 75 kDa each but is cleaved into a major single band due to autolysis in cadaver liver. The purified cadaver enzyme was a FAD-specific protein. The pH optimum was 6.6 for methylenetetrahydrofolate-NADPH oxidoreductase, 6.5 for methyltetrahydrofolate-menadione oxidoreductase, and 7.2 for NADP-menadione oxidoreductase. The Km values of human liver methylenetetrahydrofolate reductase were 17 microns for NADPH and 38 microns for methyltetrahydrofolate in the reduction of menadione, and 12 microns for NADPH in the reduction of methylenetetrahydrofolate.
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PMID:Purification and characterization of methylenetetrahydrofolate reductase from human cadaver liver. 238 27

1. Riboflavin deficiency at two levels of severity was produced in weanling rats by feeding deficient diets for 6 weeks and using neck collars to prevent coprophagy. The severity of deficiency was monitored by growth, liver flavin levels and the activation coefficient of erythrocyte glutathione oxidoreductase (NAD(P)H) (EC 1.6.4.2). Control groups, receiving the same diet with ample added riboflavin, were fed either ad lib., or were pair-fed with the deficient animals. 2. The hepatic flavoenzyme, methylenetetrahydrofolate reductase (NADPH) (EC 1.5.1.20), was very markedly affected by severe riboflavin deficiency and was significantly, but less markedly, affected by the intermediate level of deficiency. This reduction in activity was due primarily to the direct effect of the diminished supply of riboflavin, and occurred to only a small extent as a result of inanition, demonstrated by a moderate reduction in activity in the more severely food-restricted of the two pair-fed groups. Since the enzyme is assayed in the presence of its flavin cofactor, FAD, it clearly cannot be reactivated in vitro, as some other depleted flavoenzymes can. The discriminatory ability in distinguishing between severe and moderate riboflavin deficiency in vivo confers some potential advantages on this oxidoreductase as a possible index of riboflavin status. 3. The hepatic activity of another key folate-metabolizing enzyme, dihydrofolate reductase (EC 1.5.1.3), was not diminished by riboflavin deficiency in the present study. 4. The ratio, labelled 5-methyltetrahydrofolic acid:other labelled compounds derived from intraperitoneally injected pteroylglutamic acid in extracts of hepatic tissue was significantly reduced in the riboflavin-deficient groups, indicating the possibility of an effect of riboflavin deficiency on folate metabolism in vivo.
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PMID:The effect of riboflavin deficiency on methylenetetrahydrofolate reductase (NADPH) (EC 1.5.1.20) and folate metabolism in the rat. 367 70

Methylenetetrahydrofolate reductase catalyzes the reduction of methylenetetrahydrofolate to methyltetrahydrofolate. This reaction commits one carbon units to the pathways of adenosylmethionine-dependent methylation in mammalian cells. We have purified the pig liver enzyme to homogeneity and shown that it contains FAD as a non-covalently bound prosthetic group. Methylenetetrahydrofolate is not only a substrate for the reductase, but also for thymidylate synthase and for methylenetetrahydrofolate dehydrogenase. The latter reaction leads to utilization of one carbon units in de novo purine biosynthesis. A priori, one might expect that methylenetetrahydrofolate reductase activity would be modulated by cellular requirements for de novo biosynthesis of purines and pyrimidines, as well as by cellular levels of adenosylmethionine. Methylenetetrahydrofolate reductase is inhibited by dihydrofolate and its polyglutamate analogues. The Ki is 6.5 microM for dihydrofolate and decreases with each additional glutamyl residue to a minimum value of 0.013 microM for dihydropteroylhexaglutamate. The I50 for dihydropteroylhexaglutamate inhibition of reductase activity in the presence of 0.5 microM methylenetetrahydropteroylhexaglutamate is 0.07 microM. We propose that stimulation of thymidylate synthase activity (as in the replicating cell) may lead to elevations in the steady state levels of cellular dihydrofolate derivatives and to resultant inhibition of methylenetetrahydrofolate reductase activity. Thus methylenetetrahydrofolate derivatives would be spared for purine and pyrimidine biosynthesis. We have also examined the inhibition of methylenetetrahydrofolate reductase by adenosylmethionine, which serves as an allosteric effector of the enzymatic activity. Adenosylmethionine induces a slow transition in the enzyme, and leads to the inhibition of NADPH-menadione, NADPH-methylenetetrahydrofolate and methyltetrahydrofolate-menadione oxido-reductase activities.
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PMID:Modulation of methylenetetrahydrofolate reductase activity by S-adenosylmethionine and by dihydrofolate and its polyglutamate analogues. 705 69

Elevated plasma homocysteine levels are associated with increased risk for cardiovascular disease and neural tube defects in humans. Folate treatment decreases homocysteine levels and dramatically reduces the incidence of neural tube defects. The flavoprotein methylenetetrahydrofolate reductase (MTHFR) is a likely target for these actions of folate. The most common genetic cause of mildly elevated plasma homocysteine in humans is the MTHFR polymorphism A222V (base change C677-->T). The X-ray analysis of E. coli MTHFR, reported here, provides a model for the catalytic domain that is shared by all MTHFRs. This domain is a beta8alpha8 barrel that binds FAD in a novel fashion. Ala 177, corresponding to Ala 222 in human MTHFR, is near the bottom of the barrel and distant from the FAD. The mutation A177V does not affect Km or k(cat) but instead increases the propensity for bacterial MTHFR to lose its essential flavin cofactor. Folate derivatives protect wild-type and mutant E. coli enzymes against flavin loss, and protect human MTHFR and the A222V mutant against thermal inactivation, suggesting a mechanism by which folate treatment reduces homocysteine levels.
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PMID:The structure and properties of methylenetetrahydrofolate reductase from Escherichia coli suggest how folate ameliorates human hyperhomocysteinemia. 1020 87

Coenzyme F(420)-dependent methylenetetrahydromethanopterin reductase (Mer) is an enzyme of the Cl metabolism in methanogenic and sulfate reducing archaea. It is composed of identical 35-40 kDa subunits and lacks a prosthetic group. The crystal structure of Mer from Methanopyrus kandleri (kMer) revealed in one crystal form a dimeric and in another a tetrameric oligomerisation state and that from Methanobacterium thermoautotrophicum (tMer) a dimeric state. Each monomer is primarily composed of a TIM-barrel fold enlarged by three insertion regions. Insertion regions 1 and 2 contribute to intersubunit interactions. Insertion regions 2 and 3 together with the C-terminal end of the TIM-barrel core form a cleft where the binding sites of coenzyme F(420) and methylene-tetrahydromethanopterin are postulated. Close to the coenzyme F(420)-binding site lies a rarely observed non-prolyl cis-peptide bond. It is surprising that Mer is structurally most similar to a bacterial FMN-dependent luciferase which contains a non-prolyl cis-peptide bond at the equivalent position. The structure of Mer is also related to that of NADP-dependent FAD-harbouring methylenetetrahydrofolate reductase (MetF). However, Mer and MetF do not show sequence similarities although they bind related substrates and catalyze an analogous reaction.
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PMID:Structure of coenzyme F(420) dependent methylenetetrahydromethanopterin reductase from two methanogenic archaea. 1089 Dec 79

Methylenetetrahydrofolate reductase catalyzes the reduction of N(5), N(10)-methylenetetrahydrofolate to N(5)-methyltetrahydrofolate. Because this substrate is unstable and dissociates spontaneously into formaldehyde and tetrahydrofolate, the customary method to assay the catalytic activity of this enzyme has been to measure the oxidation of [14C]N(5)-methyltetrahydrofolate to N(5), N(10)-methylenetetrahydrofolate and quantify the [14C]formaldehyde that dissociates from this product. This report describes a very sensitive radioenzymatic assay that measures directly the reductive catalysis of N(5),N(10)-methylenetetrahydrofolate. The radio-labeled substrate, [14C]N(5),N(10)-methylenetetrahydrofolate, is prepared by condensation of [C(14)]formaldehyde with tetrahydrofolate and the stability of this substrate is maintained for several months by storage at -80 degrees C in a pH 9.5 buffer. Partially purified methylenetetrahydrofolate reductase from rat liver, incubated with the radio-labeled substrate and the cofactors, NADPH and FAD at pH 7. 5, generates [14C]N(5)-methyltetrahydrofolate, which is stable and partitions into the aqueous phase after the assay is terminated with dimedone and toluene. A K(m) value of 8.2 microM was obtained under conditions of increasing substrate concentration to ensure saturation kinetics. This method is simple, very sensitive and measures directly the reduction of N(5), N(10)-methylenetetrahydrofolate to N(5)-methyltetrahydrofolate, which is the physiologic catalytic pathway for methylenetetrahydrofolate reductase.
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PMID:Radioenzymatic assay for reductive catalysis of N(5)N(10)-methylenetetrahydrofolate by methylenetetrahydrofolate reductase. 1108 90

The flavoprotein Escherichia coli methylenetetrahydrofolate reductase (MTHFR) catalyzes the reduction of 5,10-methylenetetrahydrofolate (CH(2)-H(4)folate) to 5-methyltetrahydrofolate (CH(3)-H(4)folate). The X-ray crystal structure of the enzyme has revealed the amino acids at the flavin active site that are likely to be relevant to catalysis. Here, we have focused on two conserved residues, Asp 120 and Glu 28. The presence of an acidic residue (Asp 120) near the N1-C2=O position of the flavin distinguishes MTHFR from all other known flavin oxidoreductases and suggests an important function for this residue in modulating the flavin reactivity. Modeling of the CH(3)-H(4)folate product into the enzyme active site also suggests roles for Asp 120 in binding of folate and in electrostatic stabilization of the putative 5-iminium cation intermediate during catalysis. In the NADH-menadione oxidoreductase assay and in the isolated reductive half-reaction, the Asp120Asn mutant enzyme is reduced by NADH 30% more rapidly than the wild-type enzyme, which is consistent with a measured increase in the flavin midpoint potential. Compared to the wild-type enzyme, the mutant showed 150-fold decreased activity in the physiological NADH-CH(2)-H(4)folate oxidoreductase reaction and in the oxidative half-reaction involving CH(2)-H(4)folate, but the apparent K(d) for CH(2)-H(4)folate was relatively unchanged. Our results support a role for Asp 120 in catalysis of folate reduction and perhaps in stabilization of the 5-iminium cation. By analogy to thymidylate synthase, which also uses CH(2)-H(4)folate as a substrate, Glu 28 may serve directly or via water as a general acid catalyst to aid in 5-iminium cation formation. Consistent with this role, the Glu28Gln mutant was unable to catalyze the reduction of CH(2)-H(4)folate and was inactive in the physiological oxidoreductase reaction. The mutant enzyme was able to bind CH(3)-H(4)folate, but reduction of the FAD cofactor was not observed. In the NADH-menadione oxidoreductase assay, the mutant demonstrated a 240-fold decrease in activity.
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PMID:Folate activation and catalysis in methylenetetrahydrofolate reductase from Escherichia coli: roles for aspartate 120 and glutamate 28. 1137 Nov 82

As many as one-third of mutations in a gene result in the corresponding enzyme having an increased Michaelis constant, or K(m), (decreased binding affinity) for a coenzyme, resulting in a lower rate of reaction. About 50 human genetic dis-eases due to defective enzymes can be remedied or ameliorated by the administration of high doses of the vitamin component of the corresponding coenzyme, which at least partially restores enzymatic activity. Several single-nucleotide polymorphisms, in which the variant amino acid reduces coenzyme binding and thus enzymatic activity, are likely to be remediable by raising cellular concentrations of the cofactor through high-dose vitamin therapy. Some examples include the alanine-to-valine substitution at codon 222 (Ala222-->Val) [DNA: C-to-T substitution at nucleo-tide 677 (677C-->T)] in methylenetetrahydrofolate reductase (NADPH) and the cofactor FAD (in relation to cardiovascular disease, migraines, and rages), the Pro187-->Ser (DNA: 609C-->T) mutation in NAD(P):quinone oxidoreductase 1 [NAD(P)H dehy-drogenase (quinone)] and FAD (in relation to cancer), the Ala44-->Gly (DNA: 131C-->G) mutation in glucose-6-phosphate 1-dehydrogenase and NADP (in relation to favism and hemolytic anemia), and the Glu487-->Lys mutation (present in one-half of Asians) in aldehyde dehydrogenase (NAD + ) and NAD (in relation to alcohol intolerance, Alzheimer disease, and cancer).
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PMID:High-dose vitamin therapy stimulates variant enzymes with decreased coenzyme binding affinity (increased K(m)): relevance to genetic disease and polymorphisms. 1191 49


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