HUMAN ORNITHINE AMINOTRANSFERASE COMPLEXED WITH L-CANALINE AND GABACULINE - STRUCTURAL BASIS FOR SUBSTRATE RECOGNITION

Background: Ornithine aminotransferase (OAT) is a 45 kDa pyridoxal-5'- phosphate (PLP)-dependent enzyme that catalyzes the conversion of L-or nithine and 2-oxoglutarate to glutamate-F-semialdehyde and glutamic ac id, respectively. In humans, loss of OAT function causes an accumulati on of ornithine that results in gyrate atrophy of the choroid and reti na, a disease that progressively leads to blindness. In an effort to l earn more about the structural basis of this enzyme's function, we hav e determined the X-ray structures of OAT in complex with two enzyme-ac tivated suicide substrates: L-canaline, an ornithine analog, and gabac uline, an irreversible inhibitor of several related aminotransferases. Results: The structures of human OAT bound to the inhibitors gabaculi ne and L-canaline were solved to 2.3 Angstrom at 110K by difference Fo urier techniques. Both inhibitors coordinate similarly in the active s ite, binding covalently to the PLP cofactor and causing a 20 degrees r otation in the cofactor tilt relative to the ligand-free form. Aromati c-aromatic interactions occur between the bound gabaculine molecule an d active-site residues Tyr85 and Phe177, whereas Tyr55 and Arg180 prov ide specific contacts to the alpha-amino and carboxyl groups of L-cana line. Conclusions: The OAT-L-canaline complex structure implicates Tyr 55 and Arg180 as the residues involved in coordinating with the natura l substrate ornithine during normal enzyme turnover. This correlates w ell with two enzyme-inactivating point mutations associated with gyrat e atrophy, Tyr55-->His and Arg 180-->Thr, The OAT-gabaculine complex p rovides the first structural evidence that the potency of the inhibito r is due to energetically favorable aromatic interactions with residue s in the active site. This aromatic-binding mode may be relevant to st ructure-based drug design efforts against other omega-aminotransferase targets, such as GABA aminotransferase.