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Dimerisation-dependent GTPase reaction of MnmE: how potassium acts as GTPase-activating element

Andrea Scrima1, Alfred Wittinghofer

  • 1Max-Planck-Institut für Molekulare Physiologie, Dortmund, Germany.

The EMBO Journal
|June 10, 2006
PubMed

Insights

MnmE, a guanine nucleotide-binding protein, uses a novel GTP hydrolysis mechanism involving potassium-dependent G-domain dimerization. This interaction is crucial for tRNA modification and subsequent phosphoryl transfer.

Area of Science:

  • Biochemistry
  • Structural Biology
  • Molecular Biology

Background:

  • MnmE is a conserved Guanine nucleotide-binding protein essential for tRNA modification.
  • Understanding MnmE's function requires elucidating its GTP hydrolysis mechanism.

Purpose of the Study:

  • To investigate the mechanism of GTP hydrolysis by MnmE.
  • To determine the structural basis of MnmE's GTPase activity and its role in tRNA modification.

Main Methods:

  • Biochemical assays to study GTP hydrolysis.
  • X-ray crystallography to determine the structure of MnmE in complex with GDP-AlFx and potassium.
  • Site-directed mutagenesis to probe protein function.

Main Results:

  • MnmE's G-domains dimerize in a potassium-dependent manner to induce GTP hydrolysis.
  • The crystal structure reveals how subunit juxtaposition reorients the catalytic machinery, stabilizing the attacking water molecule.
  • Potassium ions act analogously to the arginine finger in Ras-RasGAP system.
  • Mutational studies demonstrate that potassium-dependent dimerization and GTP hydrolysis can be uncoupled, with domain interaction being essential for phosphoryl transfer.

Conclusions:

  • A novel GTP hydrolysis mechanism for MnmE involving potassium-dependent G-domain dimerization has been elucidated.
  • This mechanism is critical for tRNA modification, with G-domain interaction preceding phosphoryl transfer.
  • A model for G-domain juxtaposition and its role in conformational changes within the tRNA-modification center is proposed.

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