A SPECIFICITY POCKET INFERRED FROM THE CRYSTAL-STRUCTURES OF THE COMPLEXES OF ALDOSE REDUCTASE WITH THE PHARMACEUTICALLY IMPORTANT INHIBITORS TOLRESTAT AND SORBINIL

Background: Aldose reductase (AR) is an NADPH-dependent enzyme implica ted in long-term diabetic complications. Buried at the bottom of a dee p hydrophobic cleft, the NADPH coenzyme is surrounded by the conserved hydrophilic residues of the AR active site. The existence of an anion ic binding site near the NADP(+) has been determined from the structur es of the complexes of AR with citrate, cacodylate and glucose-6-phosp hate. The inhibitor zopolrestat binds to this anionic site, and in the hydrophobic cleft, after a change of conformation which opens a 'spec ificity' pocket. Results: The crystal structures of the porcine AR hol oenzyme and its complexes with the inhibitors tolrestat and sorbinil h ave been solved; these structures are important as tolrestat and sorbi nil are, pharmaceutically, the most well-studied AR inhibitors. The ac tive site of the holoenzyme was analyzed, and binding of the inhibitor s was found to involve two contact zones in the active site: first, a recognition region for hydrogen-bond accepters near the coenzyme, with three centers, including the anionic site; and second, a hydrophobic contact zone in the active-site cleft, which in the case of tolrestat includes the specificity pocket. The conformational change leading to the opening of the specificity pocket upon tolrestat binding is differ ent to the one seen upon zopolrestat binding; this pocket binds inhibi tors that are more effective against AR than against aldehyde reductas e. Conclusions: The active site of AR adapts itself to bind tightly to different inhibitors; this happens both upon binding to the inhibitor 's hydrophilic heads, and at the hydrophobic and specificity pockets o f AR, which can change their shape through different conformational ch anges of the same residues. This flexibility could explain the large v ariety of possible substrates of AR.