Carbon-13 labelling study of the coenzyme B12-dependent methylitaconate ⇌α-methyleneglutarate model rearrangement reaction and examination of potential cyclopropane intermediates

Abstract

The model rearrangement mimicking the coenzyme B12-dependent, enzyme-catalysed interconversion of α-methyleneglutaric acid with methlitaconic acid has been carried out with a carbon-13 label. This experiment demonstrates beyond doubt that the acrylate group is the migrating group in the model, as it is in the enzyme-catalysed rearrangement. Experiments designed to probe the possible occurrence of cyclopropylmethyl intermediates in the model rearrangement are also described. To this end the cis- and trans-bromomethylcyclopropanediacids (16a) and (20a) were prepared starting from the common precursor cyclopropane-1,1,2-tricarboxylic acid (13). Thus, (13) was converted into the anhydride (14) which was, in turn, reduced to the lactone (15). Opening of the lactone with HBr in acetic acid gave the desired trans-diacid (16a). For the cis-diacid (20a), the anhydride (14) was hydrolysed, esterified with diazomethane, then reduced with lithium triethylborohydride. Conversion of the resulting alcohol (19) into the bromide (20a) was effected with phosphorus tribromide. An extensive series of experiments involving treatment of the acids and their methyl and tetrahydropyranyl esters with vitamin B12S was carried out. No methylitaconic acid (3a) could be detected in any of the reaction mixtures. However, α-methyleneglutaric acid (2a) and methylglutaconic acid (21) were observed as the reaction products. The methyl toluene-p-sulphonate (22) and iodide (23) were also examined and yielded results analogous to those obtained with the bromides.