TRYPANOSOMAL NUCLEOSIDE HYDROLASE - A NOVEL MECHANISM FROM THE STRUCTURE WITH A TRANSITION-STATE INHIBITOR

Nucleoside N-ribohydrolases are targets for disruption of purine salva ge in the protozoan parasites. The structure of a trypanosomal N-riboh ydrolase in complex with a transition-state inhibitor is reported at 2 .3 Angstrom resolution, The nonspecific nucleoside hydrolase from Crit hidia fasciculata cocrystallized with p-aminophenyliminoribitol reveal s tightly bound Ca2+ as a catalytic site ligand. The complex with the transition-state inhibitor is characterized by (1) large protein confo rmational changes to create a hydrophobic leaving group site (2) C3'-e xo geometry for the inhibitor, typical of a ribooxocarbenium ion (3) s tabilization of the ribooxocarbenium analogue between the neighboring group 5'-hydroxyl and bidentate hydrogen bonds to Asn168; and (4) octa coordinate Ca2+ orients a catalytic site water and is liganded to two hydroxyls of the inhibitor. The mechanism is ribooxocarbenium stabiliz ation with weak leaving group activation and is a departure from gluco hydrolases which use paired carboxylates to achieve the transition sta te.