|Title||Structure and mechanism of the 6-oxopurine nucleosidase from Trypanosoma brucei brucei.|
|Publication Type||Journal Article|
|Year of Publication||2010|
|Authors||Vandemeulebroucke, A., C. Minici, I. Bruno, L. Muzzolini, P. Tornaghi, D. W. Parkin, W. Versées, J. Steyaert, and M. Degano|
|Date Published||2010 Oct 19|
|Keywords||Animals, Crystallography, X-Ray, Kinetics, N-Glycosyl Hydrolases, Nucleosides, Protozoan Proteins, Purinones, Structure-Activity Relationship, Trypanosoma brucei brucei|
Trypanosomes are purine-auxotrophic parasites that depend upon nucleoside hydrolase (NH) activity to salvage nitrogenous bases necessary for nucleic acid and cofactor synthesis. Nonspecific and purine-specific NHs have been widely studied, yet little is known about the 6-oxopurine-specific isozymes, although they are thought to play a primary role in the catabolism of exogenously derived nucleosides. Here, we report the first functional and structural characterization of the inosine-guanosine-specific NH from Trypanosoma brucei brucei. The enzyme shows near diffusion-limited efficiency coupled with a clear specificity for 6-oxopurine nucleosides achieved through a catalytic selection of these substrates. Pre-steady-state kinetic analysis reveals ordered product release, and a rate-limiting structural rearrangement that is associated with the release of the product, ribose. The crystal structure of this trypanosomal NH determined to 2.5 Å resolution reveals distinctive features compared to those of both purine- and pyrimidine-specific isozymes in the framework of the conserved and versatile NH fold. Nanomolar iminoribitol-based inhibitors identified in this study represent important lead compounds for the development of novel therapeutic strategies against trypanosomal diseases.