HETERONUCLEAR NMR-STUDIES OF COBALT CORRINOIDS .13. AMIDE H-1 AND N-15 NMR-STUDIES OF DIASTEREOMERISM AND THE BASE-ON BASE-OFF REACTION IN 5'-DEOXYADENOSYLCOBALT CORRINOIDS

Inverse detected H-1, N-15 HMQC spectroscopy in DMSO-d6 has been used to observe the H-1 and N-15 NMR resonances and establish the H-N connectivities of the peripheral amides of a series of five 5'-deoxyadenosylcobalt corrinoids including the beta cobalamin (i.e., coenzyme B-12) and its alpha diastereomer (in which the 5'-deoxyadenosyl ligand is in the ''lower'' axial ligand position), the diastereomeric alpha- and beta-5'-deoxyadenosylcobinamides (in which the axial 5,6-dimethylbenzimidazole nucleotide has been removed by phosphodiester hydrolysis), and the base-off analogue of the coenzyme, in which the coordinating nitrogen of the axial nucleotide is methylated. The N-15 resonances were assigned to the various side-chain amides by observation of through-space connectivities of the amide protons with other protons on the complexes via NOESY spectra in DMSO-d6. The latter could be completely assigned by analogy to the NOESY spectra of these complexes in D2O since the H-1 spectra of the 5'-deoxyadenosylcobalt corrinoids in D2O have been previously assigned. The N-15 resonances of the unsubstituted amides are found to occur in the order d, b, e, g, c, a, regardless of the nature of the axial ligands. Conversion of the beta diastereomer of the cobinamide to the alpha diestereomer causes a 2.7-ppm upfield shift of the c amide resonance suggesting that this amide interacts with the 5'-deoxyadenosyl ligand in the beta diastereomer. In the base-on beta diastereomer of the cobalamin (i.e., coenzyme B-12), the syn and anti protons of the d amide are chemical shift equivalent. Upon conversion to the base-off species, the resonance of one of these protons shifts downfield by 0.7 ppm and the N-15 resonance shifts downfield by 1.7 ppm. Amide proton chemical shift thermal gradients support the possibility of an intramolecular hydrogen bond involving the d amide in the base-on species. Based on models, this hydrogen bond is postulated to involve the noncoordinating nitrogen of the axial 5,6-dimethylbenzimidazole nucleotide as the acceptor.