Methylcobalamin synthesis

Our laboratory was the first to demonstrate the ability of thimerosal to potently inhibit methionine synthase activity in cultured human neuronal cells (Waly et al., 2004). Subsequent research has shown that inhibition results from a reduction in GSH levels and impaired methylcobalamin synthesis, under conditions where methionine synthase activity is absolutely dependent upon methylcobalamin (M. Waly, unpublished observation). The fact that autistic subjects exhibit lower GSH levels and lower methionine synthase activity lends credence to the mercury... 

Lenhert and Hodgkin (15) revealed with X-ray diffraction techniques that 5 -deoxyadenosylcobalamin (Bi2-coenzyme) contained a cobalt-carbon o-bond (Fig. 3). The discovery of this stable Co—C-tr-bond interested coordination chemists, and the search for methods of synthesizing coen-zyme-Bi2 together with analogous alkyl-cobalt corrinoids from Vitamin B12 was started. In short order the partial chemical synthesis of 5 -de-oxyadenosylcobalamin was worked out in Smith s laboratory (22), and the chemical synthesis of methylcobalamin provided a second B 12-coenzyme which was found to be active in methyl-transfer enzymes (23). A general reaction for the synthesis of alkylcorrinoids is shown in Fig. 4. 

In 1933 Challenger et al. discovered that trimethylarsine was synthesized from inorganic arsenic compounds by molds (93). Recently, McBride and Wolfe (94), have reported the synthesis of dimethylarsine from arsenate by cell extracts of the methanogenic bacterium M. O. H. Methylcobalamin is the alkylating coenzyme for this synthesis which requires reduction of arsenate to arsenite, methylation of arsenite to methylarsonic acid, reduction and methylation of methylarsonic acid to dimethylarsinic acid, and finally a four electron reduction of dimethylarsinic acid to dimethylarsine (Fig. 13). 

In methylcobalamin, X is a methyl group. This compound functions as a coenzyme for several methyltransferases, and among other things is involved in the synthesis of methionine from homocysteine (see p. 418). However, in human metabolism, in which methionine is an essential amino acid, this reaction does not occur. 

Methylbenzene halogen complex of, 3 122 iodine monochloridecomplese, 3 109 Methylchlorosilanes hydrolysis, 42 149-150, 157 pyrolysis products of, 7 356-363 Methylcobalamin, 19 151, 152 Methyl-coenzyme M reductase, 32 323-325 EPR spectra, 32 323, 325 F43 and, 32 323-324 function, 32 324-325 Methyl-CoM reductase, 32 329 Methyl cyanide, osmium carbonyl complexes, reaction, 30 198-201 Methylcyclophosphazene salts, 21 70 synthesis, 21 109... 

Fig. 1. Folate-cobalamin interaction in the synthesis of purines and pyrimidines and, therefore, of DNA. (1) In gastrointestinal mucosa cells (2) in the liver (3) in peripheral tissues. C, cobalamine DAC, desoxyadenosylcobalamine HC, hydroxy cobalamine MC, methylcobalamine F, folic acid MTHF, methyltetrahydrofolic acid THF, tetrahydrofolic acid DHF, dihydrofolic acid dUMP, deoxyuridinemonophosphate dTMP, deoxythymidine-monophosphate. (Adapted from Far-...

Vitamin B12 consists of a porphyrin-like ring with a central cobalt atom attached to a nucleotide. Various organic groups may be covalently bound to the cobalt atom, forming different cobalamins. Deoxyadenosylcobalamin and methylcobalamin are the active forms of the vitamin in humans. Cyanocobalamin and hydroxocobalamin (both available for therapeutic use) and other cobalamins found in food sources are converted to the active forms. The ultimate source of vitamin Bi2 is from microbial synthesis the vitamin is not synthesized by animals or plants. The chief dietary source of vitamin Bi2 is microbially derived vitamin B12 in meat (especially liver), eggs, and dairy products. Vitamin Bi2 is sometimes called extrinsic factor to differentiate it from intrinsic factor, a protein normally secreted by the stomach that is required for gastrointestinal uptake of dietary vitamin B12. 

The reactions catalyzed by B12 may be grouped into two classes those catalyzed by methylcobalamin and those catalyzed by cofactor B,2. The former reactions include formation of methionine from homocysteine, methanogenesis (formation of methylmercury is an important side reaction), and synthesis of acetate from carbon dioxide (82). The latter reactions include the ribonucleotide reductase reaction and a variety of isomerization reactions (82). Since dehydration and deamination have been studied quite extensively and very possibly proceed via [Pg.257]

The most notorious mercury compounds in the environment are monomethyl mercury (CH3Hg+) salts and dimethylmercury ((CH3)2Hg). The latter compound is both soluble and volatile, and the salts of the monomethylmercury cation are soluble. These compounds are produced from inorganic mercury in sediments by anaerobic bacteria through the action of methylcobalamin, a vitamin B12 analog and intermediate in the synthesis of methane ... 

Vitamin B12 is required by only two enzymes in human metabolism methionine synthetase and L-methylmalonyl-CoA mutase. Methionine synthetase has an absolute requirement for methylcobalamin and catalyzes the conversion of homocysteine to methionine (Fig. 28-5). 5-Methyltetrahydrofolate is converted to tetrahydrofolate (THF) in this reaction. This vitamin B12-catalyzed reaction is the only means by which THF can be regenerated from 5-methyltetrahydrofolate in humans. Therefore, in vitamin B12 deficiency, folic acid can become trapped in the 5-methyltetrahydrofolate form, and THF is then unavailable for conversion to other coenzyme forms required for purine, pyrimidine, and amino acid synthesis (Fig. 28-6). All folate-dependent reactions are impaired in vitamin B12 deficiency, resulting in indistinguishable hematological abnormalities in both folate and vitamin B12 deficiencies. 

The various photochemical studies of these compounds have been conducted primarily because of the interest of researchers in the synthesis, properties, and biological activity of vitamin and its derivatives. The structure of vitamin Bj2, as determined by Crowfoot-Hodgkin et al. 109), is shown in Fig. 2. It consists of cobalt in a corrin ring complexed axially by an a-S, 6-dimethylbenzimidazole nucleotide and by cyanide ion. Replacement of the axial CN by a methyl group gives methylcobalamin, and by 5 -deoxyadenosine gives coenzyme Bj2. The formal oxidation state of... 

Methylcobalamin is completely different from adenosylcobalamin because it is essentially a conduit for synthetic reactions catalyzed by methyltransferases, illustrated in Scheme 2 for the case of methionine. These reactions depend on the supemucle-ophilicity of cob(I)alamin. In one case, this species removes a methyl group from A -methyltetrahydrofolate with the formation of methylcobalamin, and then transfers this group to the acceptor homocysteine, which results in the synthesis of methionine (see Scheme 2). 

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)).

Metabolism of cobalamins in mammalian cells. Note the compartmentalization of the synthesis of the two coenzymes adenosylcobalamin (AdoCbl) synthesis occurs in mitochondria, whereas that of methylcobalamin (MeCbl) occurs in cytoplasm. TCII, Transcobalamin II OH-Cbl, hydroxocobalamin T, transport protein for TCII-OHCbl complex FH4, MeFH4, tetrahydrofolate and methyltetrahydrofolate, respectively Cbl, a cobalamin in which the ligand occupying the sixth coordination position of the cobalt is not known. The numerical superscripts adjacent to some of the cobalamins indicate the oxidation state of the cobalt ion. Cobalamins in the +1, +2, and +3 oxidation states are also known as Bn, B 2, and Bi2j, respectively. 

Inborn errors in the synthesis of adenosylcobalamin or of both adenosyl- and methylcobalamins have been described. They cause, respectively, methylmalonic aciduria alone or combined with homocystinuria (Table 38-1). These disorders respond to treatment with pharmacological doses of vitamin B12. Methylmalonic aciduria that does not respond to vitamin B12 is probably due to an abnormal methylmalonyl-CoA mutase. 

Vitamin B12 (cyanocobalamin) 3 is, in fact, not a natural product as the cyanide ligand to the cobalt ion is added during the isolation procedure. Coenzyme B12 (adenosylcobalamin) 4 and methylcobalamin 5 are the true final products of the biosynthetic pathway. Coenzyme 0,2 is the cofactor for a number of enzymic rearrangement reactions, such as that catalysed by methylmalonyl CoA mutase, and methylcobalamin is the cofactor for certain methyl transfer reactions, including the synthesis of methionine. A number of anaerobic bacteria produce related corrinoids in which the dimethylbenzimidazole moiety of the cobalamins (3 - 5) is replaced by other groups which may or may not act as ligands to the cobalt ion, such as adenine orp-cresol [12]. 

Fig. 1 GSH synthesis and methylation pathways in neuronal cells. Cysteine for GSH synthesis is provided by either uptake via EAAT3 or via transsulfuration of homocysteine (HCY), although transsulfuration is limited in neuronal cells, increasing the importance of uptake. Methionine synthase activity in neurons requires methylcobalamin (MeCbl), whose synthesis is GSH dependent. Dopamine-stimulated PLM is dependent upon methionine synthase activity. Methionine synthase activity determines levels of the methyl donor SAM and the methylation inhibitor SAH, affecting the efficiency of a large number of cellular methylation reactions.

Methionine synthase is composed of five structural domains that provide for binding of its substrate HCY, the methyl donor 5-methyItetrahydrofolate, cobal-amin, and SAM (Fig. 4). In most tissues SAM is utilized to methylate oxidized cobalamin, in conjunction with electron donation by methionine synthase reductase, thereby restoring methylcobalamin and allowing resumption of activity. This mode of reactivation is required approximately every 100-1,000 turnovers, even under strictly anaerobic laboratory conditions (Bandarian et al., 2003). Under physiological conditions, oxidation of cobalamin is undoubtedly much more common, illustrating how vitamin B12 serves as a sensor of redox status. During oxidative stress, cobalamin is more frequently oxidized and more HCY is diverted toward cysteine and GSH synthesis. 

The ratio of methylcobalamin to total vitamin derivatives of extractable B12 has been determined in liver from mice who were subjected to different types of injury (mechanical trauma, bums, and ionizing radiation) inflicted separately or in various combinations. A decrease in methylcobalamin was observed paralleling the severity of the damage. There may thus be a decreased synthesis of methycobalamin or an increased catabolism or leakage from the liver—or combination of these causes. The method used did not determine the nonextractable cobalamin, so that a disappearance into a nonextractable form could have been the cause (L9). 

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