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The McrBC restriction endonuclease assembles into a ring structure in the presence of G nucleotides

D Panne1, S A Müller, S Wirtz

  • 1Department of Microbiology and Maurice E.Müller Institute for Structural Biology, Biozentrum, Basel University, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.

The EMBO Journal
|June 19, 2001
PubMed

Insights

McrBC endonuclease from E. coli forms ring-shaped oligomers, specifically heptamers and tetradecamers, in the presence of cofactors like Mg(2+) and GTP. These structures are crucial for its DNA restriction activity.

Area of Science:

  • Molecular Biology
  • Enzymology
  • Structural Biology

Background:

  • McrBC is an Escherichia coli restriction enzyme that targets DNA with modified cytosines.
  • The mcrB gene encodes two proteins: McrB(L) (DNA-binding) and McrB(S) (lacks DNA-binding).

Purpose of the Study:

  • To investigate the quaternary structure of the McrBC endonuclease.
  • To understand the role of cofactors and subunits in McrBC structure and function.

Main Methods:

  • Gel filtration chromatography to assess oligomerization.
  • Electron microscopy (negative staining and scanning transmission) for structural visualization.
  • Mass analysis to determine particle stoichiometry.

Main Results:

  • McrB(L) and McrB(S) form high molecular weight oligomers with Mg(2+) and GTP/GDP/GTP-gamma-S.
  • Oligomerization occurs independently of DNA and GTP hydrolysis.
  • Ring-shaped particles with central channels were observed, forming heptameric rings and tetradecamers.
  • McrC presence favors the tetradecameric form, essential for DNA cleavage.

Conclusions:

  • The quaternary structure of McrBC endonuclease is cofactor-dependent.
  • McrBC forms distinct oligomeric ring structures (heptamers and tetradecamers).
  • The tetradecameric form, in conjunction with McrC, is the active endonuclease complex.

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