ACID-BASE CHEMISTRY OF ALPHA-ALKYLCOBALAMINS - DETERMINATION OF ADDITIONAL FORMATION-CONSTANTS FOR THE TUCK-IN SPECIES OF BASE-OFF COBALAMINS

Potentiometric titration of five alpha-alkylcobalamins, in which the axial organic ligand is in the 'lower' (alpha) axial position preventing coordination of the axial nucleotide, has provided the first values of the pK(a) of the conjugate acid of the pendent, but uncoordinated, benzimidazole nucleotide in cobalamins. In every case, these values are lower than the pK(a) of the conjugate acid- of the free nucleoside, alpha-ribazole, at the same temperature. This suggests that, as is the case with base-off dicyanocobalamin and base-off but benzimidazole deprotonated beta-alkylcobalamins, the free base nucleotide in the alpha-alkylcobalamins is associated with a corrin ring side chain to form what is known as the 'tuck-in' species. The data permit calculation of apparent equilibrium constants for formation of the 'tuck-in' species of the alpha-alkylcobalamins which vary from about 0.3 to 1.3, but are essentially temperature independent for a given complex. The isoenthalpic nature of this equilibrium is consistent with the principle interaction in the 'tuck-in' species being a hydrogen bond between the nucleotide B3 nitrogen and a side chain amide N-H, as is the case for dicyanocobalamin and the base-off beta-alkylcobalamins. Further evidence for formation of 'tuck-in' species in alpha-alkylcobalamins has been obtained from comparison of the C-13 NMR resonances of the pendent nucleotide of these complexes with those of the free nucleotide, alpha-ribazole-3'-phosphate, and with those of dicyanocobalamin, in which the 'tuck-in' species is well characterized. These spectral comparisons suggest that formation of the 'tuck-in' species may be accompanied by a change in conformation about the nucleotide N-glycosidic bond, and that the conformation of the nucleotide ribose moiety in the 'tuck-in' species of the alpha-alkylcobalamins may be different from that in dicyanocobalamin.