Coordination and redox state-dependent structural changes of the heme-based oxygen sensor Af GcHK associated with intraprotein signal transduction

J Biol Chem. 2017 Dec 22;292(51):20921-20935. doi: 10.1074/jbc.M117.817023. Epub 2017 Nov 1.


The heme-based oxygen sensor histidine kinase AfGcHK is part of a two-component signal transduction system in bacteria. O2 binding to the Fe(II) heme complex of its N-terminal globin domain strongly stimulates autophosphorylation at His183 in its C-terminal kinase domain. The 6-coordinate heme Fe(III)-OH- and -CN- complexes of AfGcHK are also active, but the 5-coordinate heme Fe(II) complex and the heme-free apo-form are inactive. Here, we determined the crystal structures of the isolated dimeric globin domains of the active Fe(III)-CN- and inactive 5-coordinate Fe(II) forms, revealing striking structural differences on the heme-proximal side of the globin domain. Using hydrogen/deuterium exchange coupled with mass spectrometry to characterize the conformations of the active and inactive forms of full-length AfGcHK in solution, we investigated the intramolecular signal transduction mechanisms. Major differences between the active and inactive forms were observed on the heme-proximal side (helix H5), at the dimerization interface (helices H6 and H7 and loop L7) of the globin domain and in the ATP-binding site (helices H9 and H11) of the kinase domain. Moreover, separation of the sensor and kinase domains, which deactivates catalysis, increased the solvent exposure of the globin domain-dimerization interface (helix H6) as well as the flexibility and solvent exposure of helix H11. Together, these results suggest that structural changes at the heme-proximal side, the globin domain-dimerization interface, and the ATP-binding site are important in the signal transduction mechanism of AfGcHK. We conclude that AfGcHK functions as an ensemble of molecules sampling at least two conformational states.

Keywords: bacterial protein kinase; crystal structure; globin; heme-containing oxygen sensor; histidine kinase; hydrogen-deuterium exchange; signal transduction; two component signal transduction system.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / metabolism*
  • Crystallography, X-Ray
  • Deuterium Exchange Measurement
  • Ferric Compounds / chemistry
  • Ferrous Compounds / chemistry
  • Heme / chemistry*
  • Histidine Kinase / chemistry*
  • Histidine Kinase / metabolism*
  • Mass Spectrometry
  • Models, Molecular
  • Myxococcales / metabolism
  • Oxidation-Reduction
  • Oxygen / metabolism
  • Phosphorylation
  • Protein Domains
  • Protein Structure, Quaternary
  • Signal Transduction


  • Bacterial Proteins
  • Ferric Compounds
  • Ferrous Compounds
  • Heme
  • Histidine Kinase
  • Oxygen

Associated data

  • PDB/5OHE
  • PDB/5OHF
  • PDB/2W31
  • PDB/3DGE
  • PDB/4ZVA
  • PDB/4ZVB