Cobalamins adenosylcobalamin

Figure 18 Origin of the molecular framework of cobalamin. Adenosylcobalamin is derived from aminolevulinic acid, SAM, cobalt, glutamine, threonine, ATP, NaMN, and FMN. The diagram is color coded for easy interpretation.

Metabolism and Mobilization. On entry of vitamin B 2 into the cell, considerable metaboHsm of the vitamin takes place. Co(III)cobalamin is reduced to Co(I)cobalamin, which is either methylated to form methylcobalamin or converted to adenosylcobalamin (coenzyme B>22)- The methylation requires methyl tetrahydrofolate. 

The total syntheses have yielded cobyric acid and thence cyanocobalamin. Routes to other cobalamins, eg, methylcobalamin and adenosylcobalamin, are known (76—79). One approach to such compounds involves the oxidative addition of the appropriate alkyl haUde (eg, CH I to give methylcobalamin) or tosylate (eg, 5 -A-tosyladenosine to yield adenosylcobalamine) to cobalt(I)alamine. 

Although numerous enzymatic reactions requiring vitamin B12 have been described, and 10 reactions for adenosylcobalamin alone have been identified, only three pathways in man have so far been recognized, one of which has only recently been identified (PI). Two of these require the vitamin in the adenosyl form and the other in the methyl form. These cobalamin coenzymes are formed by a complex reaction sequence which results in the formation of a covalent carbon-cobalt bond between the cobalt nucleus of the vitamin and the methyl or 5 -deoxy-5 -adenosyl ligand, with resulting coenzyme specificity. Adenosylcobalamin is required in the conversion of methylmalonate to succinate (Fig. 2), while methylcobalamin is required by a B12-dependent methionine synthetase that enables the methyl group to be transferred from 5-methyltetrahydrofolate to homocysteine to form methionine (Fig. 3). 

S-Methylmalonyl-CoA mutase (EC 5.4.99.2) is a deoxyadenoxyladen-osylcobalamin-dependent enzyme of mitochondria required to catalyze the conversion of methylmalonyl-CoA to succinyl-CoA. A decrease in the activity of methylmalonyl-CoA mutase leads to the urinary excretion of large amounts of methylmalonic acid (C22). The biochemical lesion may be at the mutase level due to an abnormality of apoenzyme protein or an inability to elaborate the required coenzyme form of vitamin B12> i.e., adenosyl-cobalamin. In rare cases the abnormality may be due to an inability to convert the d form of methylmalonyl-CoA mutase to the l form as a result of a defective racemase (EC 5.1.99.1) (Kll). In patients, the nature of the abnormality can be determined by tissue culture studies (D13) and by clinical trial, since patients with a defect in adenosylcobalamin production will show clinical improvement when treated with very large doses of vitamin B12 (Mil). 

Interaction with Adenosylcobalamin. It has been considered generally that adenosylcobalamin or its analogs binds to the apoprotein of diol dehydrase or other adenosylcobalamin-dependent enzymes almost irreversibly (4). However, we found that the holo-enzyme of diol dehydrase was resolved completely into intact apoen-zyme and adenosylcobalamin when subjected to gel filtration on a Sephadex G-25 column in the absence of K+ (9, 10). Among the inactive complexes of diol dehydrase with irreversible cobalamin inhibitors, those with cyanocobalamin and methylcobalamin also were resolved upon gel filtration on Sephadex G-25 in the absence of both K+ and substrate, yielding the apoenzyme, which was reconstitutable into the active holoenzyme (II). The enzyme-hydroxocobalamin complex, however, was not resolvable under the same conditions. The enzyme-cobalamin complexes were not resolved at all by gel filtration in the presence of both K+ and substrate. When gel filtration of the holoenzyme was carried out in the presence of K+ only, the holoen-... 

Both Components F and S were required for irreversible cleavage of the C-Co bond of adenosylcobalamin by oxygen upon aerobic incubation with the coenzyme in the absence of substrate. This suggests that activation of the C-Co bond of the coenzyme is dependent on both components. Sephadex G-25 filtration experiments showed that neither adenosylcobalamin nor cyanocobalamin was bound by the individual components, F or S. Both of them were necessary for the cobalamin binding (8). 

The ligand attached to the cobalt atom determines the activity of vitamin B12 in human enzymatic reactions. The two active coenzyme forms are methyl-cobalamin and 5 -adenosylcobalamin, the primary form of vitamin B12 in tissues. Cyanocobalamin, the therapeutic form of vitamin B12 contained in vitamin supplements, is produced by the cleavage of the unstable fink... 

Figure 1 In the above structure, R = CN denotes cyanocobalamin (CN-Cbl), whilst R = OH is hydroxocobalamin (OH-Cbl) R = 5 -deoxyadenosyl is coenzyme B12 (adenosylcobalamin, AdoCbl) and R = Me is methylcobalamin (MeCbl). By definition all cobalamins contain 5,6-dimethylbenzimidazole, which is the so-called 6th ligand to cobalt in the above structure. Substances containing the corrin ligand, i.e. the planar 14 electron p-system embracing cobalt in the above structure, are also called corrinoids.

Vitamin B12 is a mixture of cobalamins. Dietary vitamin B12 is converted to the active forms, methyl-cobalamin (mecobalamin) and adenosylcobalamin. The Average Requirement of total cobalamins in adults is 1.0 microgram/day and the Population Reference Intake is 1.4 micrograms/day. The Lowest Threshold Intake is 0.6 micrograms/day. Hydroxocobalamin (rINN vitamin Bi2a) and cyanocobalamin (rINN) have been used therapeutically. 

Structure of the cobalamin family of compounds. A through D are the four rings in the corrinoid ring system. The B ring is important for cobalamin binding to intrinsic factor. If R = -CN, the molecule is cyanocobalamin (vitamin B12) if R = 5 -deoxyadenosine, the molecule is adenosylcobalamin if R = -CH3, the molecule is methylcobalamin. Arrows pointing toward the cobalt ion represent coordinate-covalent bonds. 

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. 

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]. 

Numerous analogs of adenosylcobalamin have been tested for their ability to replace or to inhibit the action of the coenzyme in the adenosyl-cobalamin-dependent ribonucleotide reductase reaction the enzyme from L. leichmannii has been used in most of these studies. Kinetic studies have been used in most investigations of analog-enzyme interactions and thus the interpretation of data regarding the affinity of analogs for the reductase is subject to the limitations imposed on kinetic studies of a complex reaction. 

Cob(II)alamin was found to be a competitive inhibitor with respect to adenosylcobalamin of ATP reduction by the L. leichmannii enzyme. In the presence of 5 -deoxyadenosine the apparent Ki was decreased more than 10-fold adenosine was able to enhance inhibition by cob(II)-alamin to a lesser extent, but 2 -deoxyadenosine, 3 -deoxyadenosine, and 4, 5 -didehydro-5 -deoxyadenosine were much less effective. These studies indicate that cobalamin and nucleoside bind at sites that are occupied by adenosylcobalamin during catalysis and that the binding of the cobalamin moiety is greatly enhanced by the presence of a specific nucleoside (106). 

Two recently synthesized analogs of adenosylcobalamin, formycinyl-cobalamin and N6-ethenoadenosylcobalamin may be useful in studies of the mechanism of action of the coenzyme (149). The fluorescence of both nucleosides is completely quenched in the coenzyme form, but after... 

Cobalamin (B12) Adenosylcobalamin and methylcobalamin Alkyl/hydride group transfer Fig. lu... 

Figure 27 Transformation of adenosylcobyric acid into adenosylcobalamin. The pathway shown in this figure is highlighted with the aerobic pathway enzymes. The corresponding enzymes of the anaerobic pathway are shown in Table 1. It is likely that the a-ribazole is synthesized as a-ribazole phosphate and that the phosphate is removed from the final coenzyme form of cobalamin.

In S. enterica, it has recently been shown that the adenosylcobalamin 5 -phosphate is acted upon by a phosphatase called CobC. " This enzyme generates adenosylcobalamin and is therefore technically the final enzyme in cobalamin biosynthesis. ... 

---