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Science 12 May 1989:
Vol. 244. no. 4905, pp. 679 - 683
DOI: 10.1126/science.2470150
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Articles

Science, Vol 244, Issue 4905, 679-683
Copyright © 1989 by American Association for the Advancement of Science

articles

Stereochemistry of RNA cleavage by the Tetrahymena ribozyme and evidence that the chemical step is not rate-limiting

JA McSwiggen and TR Cech

Howard Hughes Medical Institute, Department of Chemistry and Biochemistry, University of Colorado, Boulder 80309-0215.

The intervening sequence of the ribosomal RNA precursor of Tetrahymena is a catalytic RNA molecule, or ribozyme. Acting as a sequence-specific endoribonuclease, it cleaves single-stranded RNA substrates with concomitant addition of guanosine. The chemistry of the reaction has now been studied by introduction of a single phosphorothioate in the substrate RNA at the cleavage site. Kinetic studies show no significant effect of this substitution on kcat (rate constant) or Km (Michaelis constant), providing evidence that some step other than the chemical step is rate-limiting. Product analysis reveals that the reaction proceeds with inversion of configuration at phosphorus, consistent with an in-line, SN2 (P) mechanism. Thus, the ribozyme reaction is in the same mechanistic category as the individual displacement reactions catalyzed by protein nucleotidyltransferases, phosphotransferases, and nucleases.


THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES:
Synthesis of phosphorothioamidites derived from 3'-thio-3'-deoxythymidine and 3'-thio-2',3'-dideoxycytidine and the automated synthesis of oligodeoxynucleotides containing a 3'-S-phosphorothiolate linkage.
G. Sabbagh, K. J. Fettes, R. Gosain, I. A. O'Neil, and R. Cosstick (2004)
Nucleic Acids Res. 32, 495-501
   Abstract »    Full Text »    PDF »
SURVEY AND SUMMARY: Recent advances in the elucidation of the mechanisms of action of ribozymes.
Y. Takagi, M. Warashina, W. J. Stec, K. Yoshinari, and K. Taira (2001)
Nucleic Acids Res. 29, 1815-1834
   Abstract »    Full Text »    PDF »
The stereochemical course of group II intron self-splicing.
R. Padgett, M Podar, S. Boulanger, and P. Perlman (1994)
Science 266, 1685-1688
   Abstract »    PDF »
Ribozymes: a distinct class of metalloenzymes.
A. Pyle (1993)
Science 261, 709-714
   Abstract »    PDF »
Aminoacyl esterase activity of the Tetrahymena ribozyme.
J. Piccirilli, T. McConnell, A. Zaug, H. Noller, and T. Cech (1992)
Science 256, 1420-1424
   Abstract »    PDF »
Structure of ribonuclease H phased at 2 A resolution by MAD analysis of the selenomethionyl protein.
W Yang, W. Hendrickson, R. Crouch, and Y Satow (1990)
Science 249, 1398-1405
   Abstract »    PDF »
A Mn2(+)-dependent ribozyme.
V Dange, R. Van Atta, and S. Hecht (1990)
Science 248, 585-588
   Abstract »    PDF »


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