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RhoC GTPase Activation Assay
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Structure-based mechanism for activation of the AAA+ GTPase McrB by the endonuclease McrC

Neha Nirwan1, Yuzuru Itoh2,3, Pratima Singh1

  • 1Division of Biology, Indian Institute of Science Education and Research, Pune, 411008, India.

Nature Communications
|July 13, 2019
PubMed

Insights

The AAA+ GTPase McrB, activated by McrC, powers DNA cleavage. Structural analysis reveals McrC bridges two McrB rings, detailing the activation mechanism and nucleotide hydrolysis pathway.

Area of Science:

  • Molecular Biology
  • Structural Biology
  • Biochemistry

Background:

  • The AAA+ GTPase McrB collaborates with endonuclease McrC to cleave DNA.
  • The precise architecture and activation mechanism of the McrBC complex were previously unknown.

Purpose of the Study:

  • To elucidate the structural basis of McrBC complex formation and activation.
  • To understand the mechanism by which McrC activates McrB for DNA cleavage.

Main Methods:

  • Reported a 3.6 Å structure of a GTPase-active, DNA-binding deficient McrBC construct.
  • Utilized structural biology techniques to visualize the complex architecture.

Main Results:

  • Revealed two hexameric McrB rings bridged by a McrC dimer.
  • Demonstrated asymmetric interaction of McrC with McrB protomers, inserting a stalk into the McrB ring pore.
  • Observed nucleotide occupancy suggesting sequential GTP hydrolysis: three pockets with 5'-guanylyl imidodiphosphate and three with GDP.
  • Identified conformational changes in key residues essential for GTP hydrolysis.

Conclusions:

  • The structure provides insights into the McrBC complex architecture and McrC-mediated activation of McrB.
  • The findings suggest a mechanism of sequential GTP hydrolysis for DNA cleavage.
  • The structural similarity to F1-ATPase highlights conserved principles in AAA+ enzyme function.

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