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Ribozymes02:47

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The term ribozyme is used for RNA that can act as an enzyme. Ribozymes are mainly found in selected viruses, bacteria, plant organelles, and lower eukaryotes. Ribozymes were first discovered in 1982 when Tom Cech’s laboratory observed Group I introns acting as enzymes. This was shortly followed by the discovery of another ribozyme, Ribonulcease P, by Sid Altman’s laboratory. Both Cech and Altman received the Nobel Prize in chemistry in 1989 for their work on ribozymes.
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Two Divalent Metal Ions and Conformational Changes Play Roles in the Hammerhead Ribozyme Cleavage Reaction.

Aamir Mir1, Ji Chen1, Kyle Robinson1

  • 1Department of Biochemistry, Purdue University , West Lafayette, Indiana 47907, United States.

Biochemistry
|September 24, 2015
PubMed
Summary
This summary is machine-generated.

The hammerhead ribozyme

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • The hammerhead ribozyme is a self-cleaving RNA found in all life forms.
  • Its catalytic mechanism, particularly the role of G12 and metal ions, remains incompletely understood.
  • Previous studies lacked crystallographic evidence of metal ion interactions at the cleavage site.

Purpose of the Study:

  • To elucidate the catalytic mechanism of the hammerhead ribozyme.
  • To investigate the role of G12 and metal ion coordination in catalysis.
  • To resolve the structural basis for hammerhead ribozyme activity.

Main Methods:

  • X-ray crystallography of wild-type and G12A mutant hammerhead ribozymes.
  • pH-dependent kinetic analysis.
  • Metal ion specificity assays.

Main Results:

  • Observed a pH-dependent conformational change around G12, indicating deprotonation.
  • Identified a Zn(2+) specificity switch in the G12A mutant, suggesting direct metal ion interaction with G12.
  • Kinetic data and metal ion binding suggest A12's minor tautomer acts as the general base in the mutant.

Conclusions:

  • A model is proposed where the hammerhead ribozyme rearranges to position two metal ions.
  • The first metal ion likely lowers the pKa of the general base (G12 or A12) and organizes the active site.
  • The second metal ion may directly participate in cleaving the RNA substrate.