Two histidine residues are essential for ribonuclease T1 activity as is the case for ribonuclease A
| Autor: | Morio Ikehara, Satoshi Nishikawa, Eiko Ohtsuka, Toshio Hakoshima, Hiroshi Morioka, Kayoko Fuchimura, Seiichi Uesugi, Hyo Joon Kim, Ken Ichi Tomita, Toshiki Tanaka |
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| Rok vydání: | 1987 |
| Předmět: | |
| Zdroj: | Biochemistry. 26:8620-8624 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi00400a019 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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