NADP-dependent enzymes. I: Conserved stereochemistry of cofactor binding.

The ubiquitous redox cofactors nicotinamide adenine dinucleotides [NAD and NADP] are very similar molecules, despite their participation in substantially different biochemical processes. NADP differs from NAD in only the presence of an additional phosphate group esterified to the 2'-hydroxyl group of the ribose at the adenine end and yet NADP is confined with few exceptions to the reactions of reductive biosynthesis, whereas NAD is used almost exclusively in oxidative degradations. The discrimination between NAD and NADP is therefore an impressive example of the power of molecular recognition by proteins. The many known tertiary structures of NADP complexes affords the possibility for an analysis of their discrimination. A systematic analysis of several crystal structures of NAD(P)-protein complexes show that: 1) the NADP coenzymes are more flexible in conformation than those of NAD; 2) although the protein-cofactor interactions are largely conserved in the NAD complexes, they are quite variable in those of NADP; and 3) in both cases the pocket around the nicotinamide moiety is substrate dependent. The conserved and variable interactions between protein and cofactors in the respective binding pockets are reported in detail. Discrimination between NAD and NADP is essentially a consequence of the overall pocket and not of a few residues. A clear fingerprint in NAD complexes is a carboxylate side chain that chelates the diol group at the ribose near the adenine, whereas in NADP complexes an arginine side chain faces the adenine plane and interacts with the phosphomonoester. The latter type of interaction might be a general feature of recognition of nucleotides by proteins. Other features such as strand-like hydrogen bonding between the NADP diphosphate moieties and the protein are also significant. The NADP binding pocket properties should prove useful in protein engineering and design.