The interaction of adenine with its binding site in rabbit muscle glyceraldehyde 3-phosphate dehydrogenase studied by fluorescence decay.

The fluorescence decay mechanism of 1, N⁶-ethenoadenosine diphosphoribose bound to rabbit muscle glyceraldehyde 3-phosphate dehydrogenase markedly differs from that of the intact coenzyme analog (εNAD⁺) bound to the same enzyme. In the latter case the fluorescence is partially quenched by interactions between the ethenoadenine ring and amino acid residues in its binding site. Binding of the nicotinamide moiety of the coenzyme thus affects the relative orientation of the adenine ring within its binding site leading to the quenching interactions. The interactions of the adenine group with its binding site induce conformational changes in the enzyme which affect the binding of additional coenzyme molecules. The nicotinamide base thus determines, indirectly, the negative cooperativity found in NAD⁺ binding.