ABSTRACT
Pyrimidine-nucleoside phosphorylase catalyzes the phosphorolytic cleavage of thymidine and uridine with equal activity. Investigation of this protein is essential for anticancer drug design. Here, the structure of this protein from Bacillus subtilis in complex with imidazole and sulfate is reported at 1.9â Å resolution, which is an improvement on the previously reported structure at 2.6â Å resolution. The localization and position of imidazole in the nucleoside-binding site reflects the possible binding of ligands that possess an imidazole ring.
Subject(s)
Bacillus subtilis/enzymology , Imidazoles/metabolism , Pyrimidine Phosphorylases/chemistry , Pyrimidine Phosphorylases/metabolism , Sulfates/metabolism , Amino Acid Sequence , Binding Sites , Catalytic Domain , Crystallization , Crystallography, X-Ray , Imidazoles/chemistry , Models, Molecular , Protein Conformation , Substrate Specificity , Sulfates/chemistryABSTRACT
Alcohol dehydrogenases belong to the oxidoreductase family and play an important role in a broad range of physiological processes. They catalyze the cofactor-dependent reversible oxidation of alcohols to the corresponding aldehydes or ketones. The NADP-dependent short-chain alcohol dehydrogenase TsAdh319 from the thermophilic archaeon Thermococcus sibiricus was overexpressed, purified and crystallized. Crystals were obtained using the hanging-drop vapour-diffusion method using 25%(w/v) polyethylene glycol 3350 pH 7.5 as precipitant. The crystals diffracted to 1.68 A resolution and belonged to space group I222, with unit-cell parameters a = 55.63, b = 83.25, c = 120.75 A.
Subject(s)
Alcohol Dehydrogenase/chemistry , Thermococcus/enzymology , Alcohol Dehydrogenase/genetics , Alcohol Dehydrogenase/isolation & purification , Crystallization , Crystallography, X-Ray , Enzyme Stability , Gene Expression , TemperatureABSTRACT
Uridine phosphorylase (UPh; EC 2.4.2.3) catalyzes the phosphorolytic cleavage of the N-glycosidic bond of uridine to form ribose 1-phosphate and uracil. This enzyme also activates pyrimidine-containing drugs, including 5-fluorouracil (5-FU). In order to better understand the mechanism of the enzyme-drug interaction, the complex of Salmonella typhimurium UPh with 5-FU was cocrystallized using the hanging-drop vapour-diffusion method at 294 K. X-ray diffraction data were collected to 2.2 A resolution. Analysis of these data revealed that the crystal belonged to space group C2, with unit-cell parameters a = 158.26, b = 93.04, c = 149.87 A, alpha = gamma = 90, beta = 90.65 degrees . The solvent content was 45.85% assuming the presence of six hexameric molecules of the complex in the unit cell.
