Ligand substitution reactions of aquacobalamin: evidence for a dissociative interchange mechanism

Abstract

The kinetics of substitution of bound H₂O in aquacobalamin (vitamin B_12a) by hydroxylamine, methyl glycinate, pyridine, 4-methylpyridine, imidazole and histamine (imidazole-4-ethanamine) was investigated as a function of ligand concentration and temperature by stopped-flow spectrophotometry at constant ionic strength (1.0 mol dm^–3) and pH. In all six cases the observed pseudo-first-order rate constants, corrected for protonation of the N-donor of the ligand, L, and the ionisation of bound H₂O in B_12a, showed saturation at high ligand concentrations. There is a compensating change in the ΔH^‡ and ΔS^‡ values of the saturating rate constant, k_sat, but there is no general isokinetic relationship for the ligands. The dependence of the value of k_sat, and, in particular, the dependence of ΔH^‡ and ΔS^‡ for this rate constant on the entering ligand, L, indicate that the rate-limiting step in the reaction is not unimolecular release of H₂O from B_12a. The results are interpreted in terms of a dissociative interchange mechanism with nucleophilic participation of L in the transition state. The compensating effect is explained on the basis of the extent of bond formation between Co and L in the transition state. The steric bulk of L, quantified using molecular mechanics techniques in terms of the cone angle subtended by co-ordinated L at Co^III, and molecular volume calculations, appears to play an important role in controlling its rate of reaction with the metal centre.