The conformation of NAD bound to dogfish lactate dehydrogenase (LDH) was reinvestigated by using an NMR expt. that allows one to exploit NOEs to det. internuclear distances between pairs of protons, without perturbation of spin-diffusion effects from other protons belonging either to the cofactor or to the binding pocket of the enzyme. The anal. indicated that the structure of bound NAD was in accord with the conformation detd. in the solid state by x-ray diffraction for the adenosine moiety, but deviated significantly from that of the nicotinamide. The NMR data indicated conformational averaging about the glycosidic bond of the nicotinamide nucleotide. In view of the strict stereospecificity of catalysis by LDH and the conformational averaging of bound NAD that was inferred from soln.-state NMR, it is suggested that LDH binds the cofactor in both syn and anti conformations, but that binding interactions in the syn conformation are not catalytically productive. [on SciFinder (R)]