Nuclear Magnetic Resonance Studies on Pyridine Dinucleotides

An assignment of the 13C nuclear magnetic resonance spectrum of NAD+ is presented based on the 13C chemical shifts of the following model compounds: protonated pyridine, benzamide, and N-methylnicotinamide. The pH dependence of the 13C spectra of NAD+ and oxidized thionicotinamide adenine dinucleotide is presented. The 13C chemical shifts of certain carbon atoms in both the adenyl and pyridyl rings are sensitive to pH. An inflection point in the pD profile occurs at a value slightly above 4 for all of these carbons. This implies that protonation of N1 in the adenine ring of NAD+ coincides with changes in the 13C chemical shifts of the pyridyl carbons. These data are interpreted to indicate the operation of a complex pH-dependent intramolecular anisotropic ring current effect in both pyridine dinucleotides.