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Monitoring ssDNA Binding to the DnaB Helicase from Helicobacter pylori by Solid-State NMR Spectroscopy.
Journal article

Monitoring ssDNA Binding to the DnaB Helicase from Helicobacter pylori by Solid-State NMR Spectroscopy.

  • Wiegand T Physical Chemistry, ETH Zurich, 8093, Zurich, Switzerland.
  • Cadalbert R Physical Chemistry, ETH Zurich, 8093, Zurich, Switzerland.
  • Gardiennet C Institut de Biologie et Chimie des Protéines, Molecular Microbiology and Structural Biochemistry, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France.
  • Timmins J Univ. Grenoble Alpes, CEA,CNRS, F-, 38044, Grenoble, France.
  • Terradot L Institut de Biologie et Chimie des Protéines, Molecular Microbiology and Structural Biochemistry, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France. laurent.terradot@ibcp.fr.
  • Böckmann A Institut de Biologie et Chimie des Protéines, Molecular Microbiology and Structural Biochemistry, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France. a.bockmann@ibcp.fr.
  • Meier BH Physical Chemistry, ETH Zurich, 8093, Zurich, Switzerland. beme@ethz.ch.
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  • 2016-10-07
Published in:
  • Angewandte Chemie (International ed. in English). - 2016
helicase
magic-angle spinning
motor proteins
solid-state NMR spectroscopy
Binding Sites
DNA, Single-Stranded
DnaB Helicases
Helicobacter pylori
Nuclear Magnetic Resonance, Biomolecular
English DnaB helicases are bacterial, ATP-driven enzymes that unwind double-stranded DNA during DNA replication. Herein, we study the sequential binding of the "non-hydrolysable" ATP analogue AMP-PNP and of single-stranded (ss) DNA to the dodecameric DnaB helicase from Helicobacter pylori using solid-state NMR. Phosphorus cross-polarization experiments monitor the binding of AMP-PNP and DNA to the helicase. 13 C chemical-shift perturbations (CSPs) are used to detect conformational changes in the protein upon binding. The helicase switches upon AMP-PNP addition into a conformation apt for ssDNA binding, and AMP-PNP is hydrolyzed and released upon binding of ssDNA. Our study sheds light on the conformational changes which are triggered by the interaction with AMP-PNP and are needed for ssDNA binding of H. pylori DnaB in vitro. They also demonstrate the level of detail solid-state NMR can provide for the characterization of protein-DNA interactions and the interplay with ATP or its analogues.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://sonar.ch/global/documents/213377
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