EPR and potentiometric studies of copper(II) binding to nicotinamide adenine dinucleotide (NAD+) in water solution

Coordination of Cu(II) by nicotinamide adenine dinucleotide (NAD+) molecule has been studied in water solutions of various pH by potentiometry and electron paramagnetic resonance (EPR) and electron spin echo (ESE) spectroscopy. Potentiometric results indicate Cu(II) coordination by protonated NAD+ at low pH and by deprotonated NAD+ at high pH. At medium pH value (around pH=7) NAD+ is not able to coordinate Cu(II) ions effectively and mainly the Cu(H2O)6 complexes exist in the studied solution. This has been confirmed by EPR results. Electronic structure of Cu(II)–NAD complex and coordination sites is determined from EPR and ESE measurements in frozen solutions (at 77K and 6K). EPR spectra exclude coordination with nitrogen atoms. Detailed analysis of EPR parameters (g||=2.420, g⊥=2.080, A||=–131×10−4cm−1 and A⊥=8×10−4cm−1) performed in terms of molecular orbital (MO) theory shows that Cu(II)NAD complex has elongated axial octahedral symmetry with a relatively strong delocalization of unpaired electron density on in-plane and axial ligands. The distortion of octahedron is analyzed using A|| vs. g|| diagram for various CuOx complexes. Electron spin echo decay modulation excludes the coordination by oxygen atoms of phosphate groups. We postulate a coordination of Cu(II) by two hydroxyl oxygen atoms of two ribose moieties of the NAD molecules and four solvated water molecules both at low and high pH values with larger elongation of the octahedron at higher pH. Cu(II) is coordinated by two nicotinamide adenine dinucleotide (NAD+) molecules in water solution of low and high pH-value. Protonation and deprotonation of the NAD+ molecule weakly affect the electronic structure of the complex as visible by electron paramagnetic resonance and electron spin echo spectroscopy. [Display omitted] ► Cu(II) binding to nicotinamide adenine dinucleotide (NAD+) in water solution. ► Protonated and deprotonated Cu(NAD+)2(H2O)4 complexes are formed at low and high pH. ► For medium pH-values mainly Cu(H2O)6 complexes exist. ► EPR and ESE exclude the coordination by nitrogen atom and oxygen of the PO4-groups.