The 13C and 1H nuclear magnetic resonance spectra and methods of their assignment for nucleotides related to dihydronicotinamide adenine dinucleotide phosphate (NADPH)

Abstract

The ¹³C n.m.r. spectra at 25·2 MHz of ribose-5-phosphate, AMP, NMN⁺, NAD⁺, NADH, NADP⁺, and NADPH have been assigned by use of a combination of techniques. In addition to established methods, such as comparisons of chemical shifts and coupling constants with those in model compounds, ionisation studies, and {¹H}–¹³C heteronuclear selective decoupling experiments, we have described some novel methods of ¹³C spectral assignments, namely (i) addition of Eu³⁺ ions to induce pseudo-contact shifts in the proton spectrum to facilitate {¹H}–¹³C decoupling experiments; (ii) the use of a graphical method of presenting results of off-resonance selective ¹H irradiation experiments; (iii) paramagnetic shift predictions for ¹³C nuclei in non-rigid molecules in the presence of Eu³⁺ ions. Difficult proton assignments can sometimes be made on the basis of connecting assigned ¹³C signals with their corresponding proton resonance signals by heteronuclear decoupling.