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3.2.2.8: ribosylpyrimidine nucleosidase

This is an abbreviated version!
For detailed information about ribosylpyrimidine nucleosidase, go to the full flat file.

Word Map on EC 3.2.2.8

Reaction

a pyrimidine nucleoside
+
H2O
=
D-ribose
 +
a pyrimidine base

Synonyms

CU-NH, cytidine-uridine nucleoside hydrolase, cytidine-uridine-preferring nucleoside hydrolase, LBRM_18_1610, More, N-ribohydrolase, N-ribosylpyrimidine nucleosidase, N-ribosylpyrimidine ribohydrolase, NH, NSH1, nucleosidase, pyrimidine, nucleoside hydrolase, pyrimidine nucleosidase, pyrimidine nucleoside hydrolase, pyrimidine-specific nucleoside hydrolase, ribonucleoside hydrolase 1, rih1, RihA, RihB, SsCU-NH, SSO0505, URH1, uridine-ribohydrolase 1, YbeK, YeiK

ECTree

     3 Hydrolases
         3.2 Glycosylases
             3.2.2 Hydrolysing N-glycosyl compounds
                3.2.2.8 ribosylpyrimidine nucleosidase

Crystallization

Crystallization on EC 3.2.2.8 - ribosylpyrimidine nucleosidase

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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method
-
hanging drop vapour diffusion method, using 100 mM Tris (pH 8.5), 200 mM NaCl, and 24% PEG 4000
molecular dynamics simulation. Both in wild-type and mutant T223Y/Q227Y, inosine binding is facilitated by interactions of the ribose moiety with active site residues and Ca2+, and pi-interactions between residues His82 and His239 and the nucleobase. The lack of observed activity toward inosine for wild-type CU-NH is explained by no residue being correctly aligned to stabilize the departing nucleobase. A hydrogen-bonding network between hypoxanthine and a general acid Asp15 is present when the two Tyr mutations are engineered into the active site. This hydrogen-bonding network is only maintained when both Tyr mutations are present due to a pi-interaction between the residues
QM/MM simulations. The relatively stronger hydrogen-bond interactions between uridine and the active-site residues Gln227 and Tyr231 play an important role in enhancing the substrate binding and thus promoting the N-glycosidic bond cleavage, in comparison with inosine. The estimated energy barrier is 30 kcal/mol for the hydrolysis of inosine and 22 kcal/mol for uridine. The uridine binding is exothermic by about 23 kcal/mol, and inosine binding by 12 kcal/mol
RihA bound to inhibitor 3,4-diaminophenyl-D-iminoribitol, hanging drop vapour diffusion method, 8 mg/ml RihA in 50 mM HEPES, pH 7.2, 150 mM NaCl is mixed with a 5:1 molar excess of 3,4-diaminophenyl-D-iminoribitol, solubilized in 50 mM HEPES, pH 7.2, and incubated at 4°C for 3 hours, the protein/inhibitor complex is mixed with an equal volume of a precipitant solution containing 25% PEG 4000, 0.1 M sodium acetate, pH 5.0, X-ray diffraction structure determination and analysis at 2.1 A resolution, molecular replacement
-
purified detagged recombinant enzyme, hanging drop vapour diffusion method, 10 mg/ml protein in 20 mM HEPES, pH 7.4, 150 mM NaCl, is mixed with precipitant solution, 25°C, equilibration versus reservoir solution containing 100 mM Tris-HCl, pH 8.5, 200 mM NaCl, 25% w/v PEG 3350, cryoprotection by 25% glycerol, X-ray diffraction structure determination and analysis at 1.7 A resolution
-
purified recombinant enzyme in complex with D-ribose, hanging drop vapour diffusion method, 10 mg/ml protein in 10 mM Tris, pH 7.0, 25 mM NaCl, and 500 mM D-ribose, is mixed with an equal volume of precipitant solution containing 24% 2-methyl-2,4-pentanediol, 0.1 M sodium acetate, pH 5.0, and 500 mM D-ribose, 20°C, 1 week, X-ray diffraction structure determination and analysis at 1.78 A resolution
to 1.53 A resolution, orthorhombic space group I222
crystal structure is determined at 1.6 A resolution. The enzyme is crystallized using the hanging drop vapor diffusion method by mixing an equal amount of protein and a precipitant solution, composed of 100 mM bicine (pH 9) and 1.5 M ammonium sulfate