The requirement for bivalent cations in formation of nicotinamide-adenine dinucleotide by nicotinamide mononucleotide adenylyltransferase of pig-liver nuclei

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Abstract
The requirement for bivalent cations in catalysis of NAD [nicotinamide adenine dinucleotide] formation from ATP [adenosine triphosphate] and NMN [nicotinamide mononucleotide] in the presence of NMN adenylyltransferase of pig-liver nuclei was studied. Rates of. NAD formation in the presence of the activating cations Cd2+, Mn2+, Mg2+, Zn2+, Co2+ and Ni2+ were approximately a linear function of heats of hydration of the corresponding ions. Ba2+, Sr2+, Ca2+, Cu2+ and Be2+ did not activate the enzyme; Be2+ inhibited the reaction in the presence of Mg2+ and, to a greater extent, in the presence of Ni2+. Michaelis constants for NAD formation, measured in a coupled assay with NMN adenylyltransferase and alcohol dehyrogenase at pH 8.0 and 25[degree], in the presence of 3 m[image] concentrations of the unvaried reactants, were 88[plus or minus]7[mu][image]-ATP, 42[plus or minus]4[mu][image]-NMN and 85 [plus or minus] 4[mu][image]-Mg2+. The results at this pH and at pH 7.5 were consistent with mechanisms in which Mg2+-ATP complex is a reactant and free ATP a competitive inhibitor. Formation of nicotinamide-hypoxanthine dinucleotide from NMN and ITP in the presence of the transf erase was also more rapid with Ni2+ and Co2+ than with Mg2+.