Methionine synthase 5-methyltetrahydrofolate

In mammals and in the majority of bacteria, cobalamin regulates DNA synthesis indirectly through its effect on a step in folate metabolism, catalyzing the synthesis of methionine from homocysteine and 5-methyltetrahydrofolate via two methyl transfer reactions. This cytoplasmic reaction is catalyzed by methionine synthase (5-methyltetrahydrofolate-homocysteine methyl-transferase), which requires methyl cobalamin (MeCbl) (253), one of the two known coenzyme forms of the complex, as its cofactor. 5 -Deoxyadenosyl cobalamin (AdoCbl) (254), the other coenzyme form of cobalamin, occurs within mitochondria. This compound is a cofactor for the enzyme methylmalonyl-CoA mutase, which is responsible for the conversion of T-methylmalonyl CoA to succinyl CoA. This reaction is involved in the metabolism of odd chain fatty acids via propionic acid, as well as amino acids isoleucine, methionine, threonine, and valine. 

N5-Methyltetrahydrofolate homocysteine methyl-transferase (= methionine synthase). This reaction is essential to restore tetrahydrofolate from N5-methyltetrahydrofolate (Fig. 2). 

Figure 45-14. Homocysteinuria and the folate trap. Vitamin 6,2 deficiency leads to inhibition of methionine synthase activity causing homocysteinuria and the trapping of folate as methyltetrahydrofolate.

When acting as a methyl donor, 5-adenosylmethionine forms homocysteine, which may be remethylated by methyltetrahydrofolate catalyzed by methionine synthase, a vitamin Bj2-dependent enzyme (Figure 45-14). The reduction of methylene-tetrahydrofolate to methyltetrahydrofolate is irreversible, and since the major source of tetrahydrofolate for tissues is methyl-tetrahydrofolate, the role of methionine synthase is vital and provides a link between the functions of folate and vitamin B,2. Impairment of methionine synthase in Bj2 deficiency results in the accumulation of methyl-tetrahydrofolate—the folate trap. There is therefore functional deficiency of folate secondary to the deficiency of vitamin B,2. 

Methionine synthase deficiency (cobalamin-E disease) produces homocystinuria without methylmalonic aciduria. This enzyme mediates the transfer of a methyl group from methyltetrahydrofolate to homocysteine to yield methionine (Fig. 40-4 reaction 4). A cobalamin group bound to the enzyme is converted to methylcobalamin prior to formation of methionine. 

The best characterized B 12-dependent methyltransferases is methionine synthase (Figure 15.11) from E. coli, which catalyses the transfer of a methyl group from methyltetrahydrofolate to homocysteine to form methionine and tetrahydrofolate. During the catalytic cycle, B12 cycles between CH3-Co(in) and Co(I). However, from time to time, Co(I) undergoes oxidative inactivation to Co(II), which requires reductive activation. During this process, the methyl donor is S-adenosylmethionine (AdoMet) and the electron donor is flavodoxin (Fid) in E. coli, or methionine synthase reductase (MSR) in humans. Methionine synthase... 

This cobalamin-dependent enzyme [EC 2.1.1.13], also known as methionine synthase and tetrahydropteroyl-glutamate methyltransferase, catalyzes the reaction of 5-methyltetrahydrofolate with L-homocysteine to produce tetrahydrofolate and L-methionine. Interestingly, the bacterial enzyme is reported to require 5-adenosyl-L-methionine and FADH2. See also Tetrahydropteroyl-triglutamate Methyltransferase... 

The loss of a methyl group from AdoMet in each of the reactions yields S-ad-enosylhomocysteine (AdoHcy) and this is subsequently hydrolysed to adenosine and Hey by AdoHcy-hydrolase. Hey sits at a metabolic branch point and can be remethylated to methionine by way of two reactions. One is the 5-methyltetrahydrofo-late dependent reaction catalysed by methionine synthase, which itself is reductively methylated by cobalamin (vitamin B12) and AdoMet, requiring methionine synthase reductase. 5-Methyltetrahydrofolate is generated from 5,10-methylenetetrahydrofo-late (MTHF) by MTHF reductase. The second remethylation reaction is catalysed by betaine methyltransferase, which is restricted to the liver, kidney and brain, while methionine synthase is widely distributed. 

A5-Methyltetrahydrofolate is the methyl-group donor substrate for methionine synthase, which catalyzes the transfer of the five-methyl group to the sulfhydryl group of homocysteine. This and selected reactions of the other folate derivatives are outlined in figure 10.15, which emphasizes the important role tetrahydrofolate plays in nucleic acid biosynthesis by serving as the immediate source of one-carbon units in purine and pyrimidine biosynthesis. 

Vitamin B12 is a biologically active corrinoid, a group of cobalt-containing compounds with macrocyclic pyrrol rings. Vitamin B12 functions as a cofactor for two enzymes, methionine synthase and L-methylmalonyl coenzyme A (CoA) mutase. Methionine synthase requires methylcobalamin for the methyl transfer from methyltetrahydrofolate to homocysteine to form methionine tetrahy-drofolate. L-methylmalonyl-CoA mutase requires adenosylcobalamin to convert L-methylmalonyl-CoA to succinyl-CoA in an isomerization reaction. An inadequate supply of vitamin B12 results in neuropathy, megaloblastic anemia, and gastrointestinal symptoms (Baik and Russell, 1999). 

In contrast to coenzyme Bi2, where the alkyl moiety serves purely in a catalytic role, the alkyl group of methyl cobamides (MeCba s) is utilized as a reagent by MeCba-dependent enzymes it is only the cobamide portion of the coenzyme that is catalytic. The cobamide-dependent methyl transferases have been reviewed [11,24-27,165], Three cobamide-dependent methyl transferases have been studied in some cases, more than one protein is required. The Bi2 proteins include methionine synthase (officially called 5-methyltetrahydrofolate-L-homocysteine-S-methyltransferase [HCM] EC 2.1.1.13) MeCba-dependent enzyme from Meth-anosarcina barkeri (MT 0 and the corrinoid/Fe-S protein from Clostridium ther-moaceticum. 

The cobalt center in MeCbl, one of the two important B12 coenzymes, is clearly involved in key steps in catalytic methyl transfer processes. Here, the Co center cycles between Co(I) and Co(III)CH3. In methionine synthase, the proposed mechanism involves direct nucleophilic attack on the C of the Co(III)CH3 group. In model reactions, the thiolate most frequently simply binds tram to the alkyl group to give a product recently established by an x-ray study of a model system. The protein may block access to the Co, thus preventing this reaction common in models. It is likely that the reactive form of the bound cofactor is five-coordinate in the key point in the catalytic cycle. This reactive form will lead to a four-coordinate Co(I) species. The axial coordination of the cofactor by a protein imidazole allows for a finer tuning of the Cbl chemistry and may permit control of the coordination number. Thus, recoordination of Co in the Co(I) state may facilitate attack on methyltetrahydrofolate and re-formation of Co(III)CH3. 

In the folate cycle, which is linked to the methionine cycle, homocysteine is remethylated to methionine by the vitamin B -dependent enzyme methionine synthase (MS), thereby completing the cycle. 5-Methyltetrahydrofolate (CH3-THF) acts as a methyl donor in this reaction, which produces methionine and tetrahydrofolate (THF). 

Figure 21-2. Metabolism of homocysteine. BHMT, betaineihomocysteine methyl-transferase CBS, cystathionine P-synthase Cob, cobalamin CTH, cystathionine y-lyase DHF, dihydrofolate DMG, dimethylglycine FAD, flavin adenine dinucleotide MAT, methionine adenosyltransferase 5-MTHF, 5-methyltetrahydrofolate 5,10-MTHF, 5,10-methylenetetrahydrofolate MTHFR, methylenetetrahydrofolate reductase MS, methionine synthase MTRR, methionine synthase reductase MTs, methyl transferases PLE pyridoxal phosphate SAH, S-adenosylhomocysteine SAHH, SAH hydrolase SAM, 5-adenosylmethionine SHMT, serine hydroxymethyltransferase THF, tetrahydrofolate Zn, zinc.

Figure 28-5. The reaction catalyzed by methionine synthase, a vitamin B12-requiring enzyme. In this reaction, homocystine is converted to methionine, with the simultaneous production of tetrahydrofolate (THF) from 5-methyltetrahydrofolate.Methionine can then be converted to 5-adenosyhnethionine (SAM), the universal methyl-group donor.

Fig. 6 Methyl trap hypothesis 5,10-Methylenetetrahydrofolate is reduced to 5-methyltetiahy-drofolate in an irreversible reaction. When vitamin Bn is deficient, methyl groups are trapped as 5-methyltetrahydrofolate, resulting in decreased substrates for DNA synthesis and neural lipid methylation. MTHFR, methylenetetrahydrofolate reductase DHFR, dihydrofolate reductase MS, Methionine synthase TS, thymidylate synthase SAM, S-adenosyl-methionine dUMP, deoxyuridine 5 -monophosphate dTTP, deoxythymidine 5 -monophosphate...

Scheme 2 Two-step synthesis of methionine from homocysteine catalyzed by methionine synthase and using methylcobalamin (MeCbl) derived from N -methyltetrahydrofolate and cob(l)alamin (Cbl(l)).

Taurog RE, Matthews RG. Activation of methyltetrahydrofolate by cobalamin-independent methionine synthase. Biochemistry 2006 45 5092-5102. 

Methionine can be regenerated by the transfer of a methyl group to homocysteine fromTV -methyltetrahydrofolate, a reaction catalyzed by methionine synthase (also known as homocysteine methyltransferase). 

The second reaction requiring vitamin B12 catalyzes the conversion of homocysteine to methionine and is catalyzed by methionine synthase. This reaction results in the transfer of the methyl group from N -methyltetrahydrofolate to hydroxycobalamin generating tetrahydrofolate and methylcobalamin during the process of the conversion. 

The inability to absorb Vitamin B12 occms in pernicious anemia. In pernicious anemia intrinsic factor is missing. The anemia results from impaired DNA synthesis due to a block in purine and thymidine biosynthesis. The block in nucleotide biosynthesis is a consequence of the effect of vitamin B12 on folate metabolism. When vitamin B-12 is deficient essentially all of the folate becomes trapped as the N -methyltetrahydrofolate derivative as a result of the loss of functional methionine synthase. This trapping prevents the synthesis of other tetrahydrofolate derivatives. required for the purine and thymidine nucleotide biosynthesis pathways. 

Methylation is the addition of a carbon atom to a molecule, usually causing a change in the function of the methylated molecule. For example, methylation of the neurotransmitter dopamine by catechol-O-methyltransferase renders it inactive. With only two exceptions, 5-adenosylmethionine (SAM), an activated form of the essential amino acid methionine, is the methyl donor for each of the more than 150 methylation reactions, which regulate a large number of cellular functions. One exception is methylation of homocysteine (HCY) to methionine by the cobalamin (vitamin Bi2)-dependent enzyme methionine synthase, which utilizes 5-methyltetrahydrofolate (methylfolate) as the methyl donor, serving to complete the methionine cycle of methylation, as illustrated in Fig. 1 (lower right). Notably, HCY formation from S-adenosylhomocysteine (SAH) is reversible and, as a result, any decrease in methionine synthase activity will be reflected as an increase in both HCY and SAH. This is significant because SAH interferes with SAM-dependent methylation reactions, and a decrease in methionine synthase activity will decrease all of these reactions. Clearly methionine synthase exerts a powerful influence over cell function via its control over methylation. 

C. Methionine can be synthesized by the methylation of homocysteine by the enzyme methionine synthase, which requires the participation of vitamin and 5-methyltetrahydrofolate. It is actually the homocysteine component of methionine that is required, since this reaction has the capacity to synthesize enough methionine. However, there is no good dietary source of homocysteine. This conversion of homocysteine to methionine also serves the purpose of making THF available for other biosynthetic reactions. 

B. Excess folate, by overwhelming the folate pool trapped as N -methyltetrahydrofolate, can allow for formation of AI, A4°-methyl-enetetrahydrofolate which is required for the thymidylate synthase reaction for DNA synthesis and red blood cell formation. Folate is not recognized as a methyl donor by methionine synthase. Folate does not inhibit destruction of erythrocytes. Cobalamin is an important critical vitamin not synthesized by humans. 

There are two classes of methionine synthases, the cobalamin-dependent and cobalamin-independent enzymes. Both synthases catalyze the same reaction, Equation (20), the methylation of (5)-homocysteine (homocysteine) by -methyltetrahydrofolate (methyltetrahydrofolate). 

One form of methionine synthase common in bacteria uses lV -methyltetrahydrofolate as a methyl donor. Another form of the enzyme present in some bacteria and mammals uses A/ -methyltetrahydro-folate, but the methyl group is first transferred to cobalamin, derived from coenzyme B12, to form methylcobalamin as the methyl donor in methionine formation. This reaction and the rearrangement of L-methyl-malonyl-CoA to succinyl-CoA (see Box 17-2, Fig. la) are the only known coenzyme Bi2-dependent reactions in mammals. In cases of vitamin B12 deficiency, some symptoms can be alleviated by administering not only vitamin B12 but folate. As noted above, the methyl group of methylcobalamin is derived from W -methyltetrahy-drofolate. Because the reaction converting the methylene form to the 7V -methyl form of tetrahydrofo-... 

Most one-carbon transfers are promoted by one of three cofactors biotin, tetrahydrofolate, or S-adenosyhnethionine (Chapter 18). S-Adenosylmethionine is generally used as a methyl group donor the transfer potential of the methyl group in 7V -methyltetrahydrofolate is insufficient for most bios3Tithetic reactions. However, one example of the use of 7V -methyltetrahydrofolate in methyl group transfer is in methionine formation by the methionine synthase reaction (step of Fig. 22-15) methionine is the immediate precursor of... 

Because methionine synthase is the only mammalian enzyme known to act on 5-methyltetrahydrofolate, the decreased intracellular activity of this enzyme causes 5-methyltetrahydrofolate to accumulate, at the expense of depleted pools of the other tetrahydrofolate coenzymes. Thus, even though total folate levels may seem ample, there is a functional folate deficiency, with insufficient levels of the formyl and methylene derivatives needed for synthesis of nucleic acid precursors. 

Methionine synthase (MetH) of E. coli represents the most thoroughly studied B12-dependent methyl transferase and is one of the essential roles of B12 in mammalian metabolism [125,153,154]. It is a modular enzyme containing separate binding domains for homocysteine, N -methyltetrahydrofolate, S-adenosyl-methionine (SAM) and the Bi2-cofactor [125,153-155]. The B12-binding domain in its different oxidation states must interact punctually and specifically with each of the other three domains The Co(I) form with the N -methyltetrahydrofolate binding domain, the Co(II) form with the SAM binding domain, and the CH3 - Co (III) form with the homocysteine binding domain [153,155]. 

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