Biochemical and Structural Investigations on Kynurenine Aminotransferase II: An Example of Conformation-Driven Species-Specific Inhibition?

Valentina      Casazza; Franca      Rossi; Menico      Rizzi

Abstract

Kynurenic acid (KYNA), one of the metabolites belonging to the kynurenine pathway, has been described as an important neuroprotective compound, its unbalancing being associated with several pathological conditions. In human brain, the majority of KYNA production is sustained by kynurenine aminotransferase II (KAT II). A selective KAT II inhibitor would be an important pharmacological tool, since it would reduce KYNA formation without causing complete depletion of this neuroprotector. (S)-(4)-(ethylsulfonyl)benzoylalanine (S-ESBA), described as a potent and selective inhibitor of rat KAT II, is unfortunately ineffective towards the human enzyme although the two orthologs share a remarkably high degree of sequence identity. We investigated the molecular basis for this intriguing species-specificity by adopting a site-directed mutagenesis and structural approach. We propose that the source of the inhibitor specificity toward the rat enzyme could reside on S-ESBA interaction/interference with a flexible loop that controls ligand admission to the active site by a classical induced-fit mechanism. Our data further highlights that even in case of highly conserved molecular targets, the flexibility of catalytically important structural elements can have a significant impact on the selectivity of inhibitor action.

Keywords: Kynurenic acid, kynurenine aminotransferase II, PLP-enzymes, crystal structure, conformational flexibility, drug design, cognition enhancing drugs, drug selectivity, S-ESBA, ligand, L-tryptophan, mammals, kynurenines, neuroactive, NMDA receptor, 7 nicotinic receptor, schizophrenia, Neurochemistry studies, dopamine, PLP-KYN, ketoacid, PMP, KAT II, mAAT, L-KYN, QUIN, hKAT II, Asp230, Tyr233, Tyr74, Arg399, isozymes, enzymes, PCR-based, homogeneity, X-ray, DNA sequencing, oligomeric, Bradford assay, albumine, Protein aliquots, UV/HPLC, ESRF, MOSFLM, SCALA, 3OH-KYN, ketoglutarate, IC50 value, catalysis, kinetics, mutant, wild-type, co-crystallization, stabilization, N-terminal