Two histidine residues are essential for ribonuclease T1 activity as is the case for ribonuclease A.
Ribonuclease T1 (RNase T1, EC 3.1.27.3) is a guanosine-specific ribonuclease that cleaves the 3',5'-phosphodiester linkage of single-stranded RNA. It is assumed that the reaction is generated by concerted acid-base catalysis between residues Glu-58 and His-92 or His-40. From the results of chemical modification and NMR studies, it appeared that the residue Glu-58 was indispensable for nucleolytic activity. However, we have recently demonstrated that Glu-58 is an important but not an essential residue for catalytic activity, using the methods of genetic engineering to change Glu-58 to Gln-58 etc [Nishikawa, S., Morioka, H., Fuchimura, K., Tanaka, T., Uesugi, S., Ohtsuka, E., & Ikehara, M. (1986) Biochem. Biophys. Res. Commun. 138, 789-794]. In the present paper, we report that mutants of RNase T1 with residue Ala-40 or Ala-92 have almost no activity, while mutants that contain Ala-58 retain considerable activity. These results show that the two histidine residues, His-40 and His-92, but not Glu-58, are indispensable for the catalytic activity of the enzyme. We propose a revised reaction mechanism in which two histidine residues play a major role, as they do in the case of RNase A.
D001236:Aspergillus oryzae / Q000201:enzymology / Q000235:genetics; D001483:Base Sequence; D003062:Codon; D004722:Endoribonucleases / Q000378:metabolism*; D005796:Genes; D005800:Genes, Fungal; D006639:Histidine; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D006163:Ribonuclease T1 / Q000235:genetics / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000378:metabolism*
D003062:Codon; D006639:Histidine; D004722:Endoribonucleases; D006163:Ribonuclease T1; D012259:Ribonuclease, Pancreatic