title: Binding of nicotinamide-adenine dinucleotides to diphtheria toxin.
abstract: 1. Changes in protein fluorescence have been utilized in determining the stoicheiometry and dissociation constants of the complexes of diphtheria toxin with NADH(2), NAD, NADPH(2) and NADP. 2. The binding stoicheiometry is 2moles of NADH(2) and 1mole of NADPH(2)/mole of diphtheria toxin. The binding sites for NADH(2) appear to be equivalent and independent. 3. The toxin shows a higher affinity for the reduced than for the oxidized forms of the nucleotides. 4. Dissociation constants at 0.01I, pH7 and 25 degrees are 0.7x10(-6)m for NADH(2) and 0.45x10(-6)m for NADPH(2). Dissociation constants increase with increasing ionic strength, indicating that the binding is mainly electrostatic. 5. Bound NADH(2) and NADPH(2) may be activated to fluoresce by the transfer of energy from the excited aromatic amino acids of the toxin. Activation and emission spectra of bound and free nucleotides are compared. 6. Since NAD and NADH(2) are cofactors specifically required for the inhibition of protein synthesis by diphtheria toxin, the possible role of toxin-nucleotide complexes is discussed in this regard.
mesh_terms: D001665:Binding Sites; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004167:Diphtheria Toxin; D005470:Fluorometry; D007477:Ions; D009243:NAD; D009249:NADP; D010084:Oxidation-Reduction; D013696:Temperature
chemical_list: D004167:Diphtheria Toxin; D007477:Ions; D009243:NAD; D009249:NADP