title: [Studies on the reaction mechanism of a ribonuclease II from Aspergillus oryzae (author's transl)].
abstract: Ribonuclease T2 was isolated from an Aspergillus oryzae extract. In order to define the substrate specificity, the hydrolysis of a series of 2',3'-cyclic nucleotides was measured semiquantitatively. Modifications in all positions of the bases are tolerated, as long as the base stays in the anti conformation or has a chance to return to it; bulky substituents at N-3 of the pyrimidine base lower the rate. So far the conclusion seems justified that the enzyme does not react with the substrates by specific bonds to the base, but rather by hydrophobic binding. The conformation specificity and the pH dependence of the activity support this hypothesis. The pH optima with substrates which may be positively or negatively charged are shifted to pH values at which the substrates are uncharged. This strongly indicates a hydrophobic type of interaction between base and enzyme. From the pH dependence of the kinetic parameters Km and k+2, an enzyme group with a pK of 7 (probably histidine) can be postulated. This group should interact in the protonated form with the phosphate anion. Another B.HB-system (probably two carboxylate groups) seems to be involved in the catalysis step, performing the base catalysis at the 2'-OH group and the proton catalysis at the phosphate oxygen simultaneously.
mesh_terms: D001230:Aspergillus; D001665:Binding Sites; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D009712:Nucleotides, Cyclic / Q000032:analysis; D011487:Protein Conformation; D011684:Purine Nucleosides / Q000032:analysis; D011741:Pyrimidine Nucleosides / Q000032:analysis; D012260:Ribonucleases / Q000302:isolation & purification / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D009712:Nucleotides, Cyclic; D011684:Purine Nucleosides; D011741:Pyrimidine Nucleosides; D012260:Ribonucleases
vernacular_title: Untersuchungen zum Reaktionsmechanismus der Ribonuclease II aus Aspergillus oryzae (Ribonuclease T2)