The mechamism of action of vitamin B12

A novel rearrangement reaction is introduced as a model for the rearrangement of methylitaconic acid (III) to α-methyleneglutaric acid (IV), one of three enzyme catalyzed, coenzyme B₁₂-dependent, carbonskeleton rearrangements whose mechanism has been a source of puzzlement for many years. The key feature of the new model is the direct attachment of the substrate, methylitaconic acid, to the cobalt atom of vitamin B₁₂ This was accomplished by reacting butadiene-2,3-decarboxylic acid with hydrobromic acid generating bromomethylitaconic acid (VIII). Use of two moles of hydrobromic acid yielded bis-2,3-(bromomethyl)succinic acid (IX). Reaction of the monobromide VIII with vitamin B_12s did not yield the desired carbon-cobalt bonded adduct. Instead, the lactone ηa- methylene-γbutyrolactone-β-carboxylic acid (X) was formed. Accordingly, the ester, dimethyl bromomethylitaconate (XIa), was reacted with vitamin B_12s and yielded the carbon-cobalt bonded adduct XIIa. Bis-trimethylsilyl bromomethylitaconate did not yield an adduct when reacted with vitamin B_12s, but bis-tetrahydropyranyl bromomethylitaconate (XIb) did yield the adduct XIIb. The ester cobalamin XIIb undergoes spontaneous decomposition at room temperature, in aqueous solution, at pH 8 and in the dark - biochemically ideal circumstances - yielding a mixture of butadiene-2,3-decarboxylic acid (VII), methylitaconic acid (III) and α- methyleneglutaric acid (IV). The presence of the latter indicates that a skeletal change has taken place in a way which mimics the enzymatic reaction. This is the first non-enzymic model in this carbon-skeleton rearrangement series. The methyl ester cobalamin XIIa was stable in the dark but did decompose on irradiation with a sunlamp to butadiene-2,3decarboxylic acid (VII) and methylitaconic acid (III). No α-methyleneglutaric acid IV was observed in the latter reaction.Authentic methylitaconic acid (III) was prepared by alkylation of triethyl prop-2-ene-l,l,2-tricarboxylate (XIII) with methyl iodide followed by hydrolysis and decarboxylation. The lactone X and lactone α-methyl-γ-butyrolactone-β-carboxylic acid (XVI) were prepared by condensing the triester XIII with formaldehyde, hydrolyzing the lactone diester XV to the lactone X and hydrogenating to the saturated lactone XVI.