Chemical and enzymatic probing of effector-mediated changes in the conformation of a maxizyme

Jing Min Zhou, Yuka Nakamatsu, Tomoko Kuwabara, Masaki Warashina, Yoshiyuki Tanaka, Koichi Yoshinari, Kazunari Taira

Research output: Contribution to journalArticlepeer-review

11 Citations (Scopus)

Abstract

The protein encoded by chimeric BCR-ABL mRNA causes chronic myelogenous leukemia (CML). We showed previously that a novel allosterically controllable ribozyme, of the type known as a maxizyme, can cleave this mRNA, with high specificity and high-level activity in vivo. We designed the maxizyme in such a way that it was able to form an active core with which to capture the catalytically indispensable Mg2+ ions only in the presence of the BCR-ABL mRNA junction. In order to probe the putative conformational changes, we used a weakly alkaline solution (pH 9.2) in the presence of 25 mM Mg2+ ions to hydrolyze differentially phosphodiester bonds that were located in different environments. Phosphodiester bonds in single-stranded regions were clearly more susceptible to attack by alkali than those within a double-stranded helix. As indicated by earlier data obtained in vivo, our results demonstrated that the active conformation was achieved only in the presence of the junction within the chimeric BCR-ABL mRNA. Moreover, we demonstrated that the use of mild alkaline solutions to probe RNA structures is very informative. Copyright (C) 2000 Elsevier Science Inc.

Original languageEnglish
Pages (from-to)261-268
Number of pages8
JournalJournal of Inorganic Biochemistry
Volume78
Issue number4
DOIs
Publication statusPublished - 2000 Mar 31

Keywords

  • Alkaline hydrolysis
  • Chemical probing
  • Chronic myelogenous leukemia
  • Maxizyme
  • RNase T1

ASJC Scopus subject areas

  • Biochemistry
  • Inorganic Chemistry

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