Catalysis by nucleoside hydrolases

Abstract

Nucleoside hydrolases cleave the N-glycosidic bond of ribonucleosides. Because of their vital role in the protozoan purine salvage pathway, nucleoside hydrolases from parasitic protozoa in particular have been studied extensively by X-ray crystallography, kinetic methods and site-directed mutagenesis. An elaborate network of conserved interactions between the metalloenzyme and the ribose enables steric and electrostatic stabilisation of the oxocarbenium-ion-like transition state. Activation of the leaving group by protonation before the formation of the transition state is a recurring catalytic strategy of enzymes that cleave N-glycosidic bonds. However, the mechanisms underlying leaving group activation are still the subject of debate for the nucleoside hydrolases.

Design of inhibitors for nucleoside hydrolase from Leishmania donovani using molecular dynamics studies

“…In the active site of CfNH, however, Glu232 is replaced by Arg233. 13,15,16,18 Superimposing the residues of the active sites from our model and the templates, it can be initially inferred, based on the alignment in Figure 1 and the picture of the active site from CfNH reported by Versées and Steyaert, 18 that, in LdNH and LmNH, residues Asp10, Asp14, Asp15, Asn39, Thr126, Asn160, Glu166, Asn168 and Asp242, would be the ones that stabilize the binding of the ribose portion of the nucleosides, while the hydrophobic pocket of the nucleic base would be formed by residues Ile81, His82, Asn160, Phe167, Tyr225, Tyr229 and His241. Also, residues Asp10, Asp15, Thr126 and Asp242 would be the ones binding the Ca 2+ ion.…”

“…[9][10][11] Parasitic nucleoside hydrolases (NH) have been extensively studied as potential targets for chemotherapy. [12][13][14][15][16][17][18] These enzymes are widely distributed in nature and have already been found in bacteria, 19,20 yeast, 21 protozoa 16,22 and insects. 23 Their encoding genes are present in plants, amphibians and fish but, surprisingly, no NH activity or their encoding genes have been detected in mammals until the present moment.…”

“…23 Their encoding genes are present in plants, amphibians and fish but, surprisingly, no NH activity or their encoding genes have been detected in mammals until the present moment. 18 This fact turns NH into ideal targets for selective chemotherapy for parasitic and bacterial infections.…”

“…[14][15][16] The analysis of the crystallographic structure of NH from Crithidia fasciculata complexed with p-aminophenyliminoribitol (pAPIR) shows that each NH monomer is functional and contains a narrow and deep active site with a strongly bound Ca 2+ ion. 18,[26][27][28] This cation is octacoordinated by a net of interactions involving the O atoms of the side chains of residues Asp10, Asp15, Asp242, the carbonyl oxygen of the main chain of Thr126, and three water molecules. In the enzyme-substrate complex, two of the water molecules are replaced by the 2' and 3' hydroxyl groups of the substrate ribose.…”

“…The specificity of NH for the ribose fraction is established by a net of conserved interactions involving the 2', 3' and 5' hydroxyl groups of the sugar, the conserved residues Asp14, Asn160, Glu166, Asn168 and Asp242 and the Ca 2+ ion. 18 Nonspecific NH catalyze the hydrolysis of purine and pirimidine nucleosides, as well as the hydrolysis of p-nitrophenyl--D-ribofuranoside. In contrast, specific NH prefer purine nucleosides, being poor catalysts for pirimidine nucleosides and p-nitrophenyl--D-ribofuranoside hydrolysis.…”