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Physicochemical Characterization of the Catalytic Unit of Hammerhead Ribozyme and Its Relationship with the Catalytic Activity
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SYSNO ASEP 0578599 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve SCOPUS Title Physicochemical Characterization of the Catalytic Unit of Hammerhead Ribozyme and Its Relationship with the Catalytic Activity Author(s) Tanaka, Y. (JP)
Yamanaka, D. (JP)
Morioka, S. (JP)
Yamaguchi, T. (JP)
Morikawa, M. (JP)
Kodama, T. S. (JP)
Sychrovský, Vladimír (UOCHB-X) RID, ORCID
Kojima, C. (JP)
Hattori, Y. (JP)Source Title Biophysica. - : MDPI AG
Roč. 2, č. 3 (2022), s. 221-239Number of pages 19 s. Language eng - English Country CH - Switzerland Keywords hammerhead ribozyme ; mechanism ; pKa ; pH-titration ; NMR spectroscopy ; G12 ; G10.1 OECD category Biophysics Method of publishing Open access Institutional support UOCHB-X - RVO:61388963 EID SCOPUS 85176554552 DOI 10.3390/biophysica2030022 Annotation The catalytic mechanism of hammerhead ribozymes (HHRzs) attracted great attention in relation to the chemical origin of life. However, the basicity (pKa) of the catalytic sites of HHRzs has not been studied so far. As a result, the investigation of the currently assumed mechanism from an experimentally derived pKa value has been impossible. In HHRzs, there exists a highly functionalized structural unit (A9-G10.1 site) with a catalytic residue (G12) for the nucleophile activation and metal ion-binding residue (G10.1). As inferred from this fact, there might be a possibility that HHRzs may utilize specific functions of the A9-G10.1 motif for the catalytic reaction. Therefore, here we studied the basicity of G12/G10.1-corresponding residues using RNA duplexes including the A9-G10.1 motif without other conserved residues of HHRzs. From the pH-titration experiments with NMR spectra, we have obtained the intrinsic basicity of the G12/G10.1-corresponding residues in the motif, with pKa > 11.5 (N1 of G12) and pKa 4.5 (N7 of G10.1) for the first time. Based on the derived irregular basicity, their correlation with a catalytic activity and a metal affinity were investigated. In total, the derived thermodynamic properties are an intrinsic nature of the exclusive catalytic unit of HHRzs, which will be an outstanding pivot point for the mechanistic analyses. Workplace Institute of Organic Chemistry and Biochemistry Contact asep@uochb.cas.cz ; Kateřina Šperková, Tel.: 232 002 584 ; Jana Procházková, Tel.: 220 183 418 Year of Publishing 2024 Electronic address https://doi.org/10.3390/biophysica2030022
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