Disruption of the dimerization interface of the sensing domain in the dimeric heme-based oxygen sensor Af GcHK abolishes bacterial signal transduction

DOI: 10.1074/jbc.RA119.011574

Authors: Tereza Skalova (Institute of Biotechnology of the Czech Academy of Sciences) , Alzbeta Lengalova (Charles University) , Jan Dohnalek (Institute of Biotechnology of the Czech Academy of Sciences) , Karl Harlos (University of Oxford) , Peter Mihalcin (Charles University) , Petr Kolenko (Czech Technical University in Prague) , Martin Stranava (Charles University) , Jan Blaha (Charles University) , Toru Shimizu (Charles University) , Markéta Martínková (Charles University)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry

State: Published (Approved)
Published: December 2019
Diamond Proposal Number(s): 10627

Abstract: The heme-based oxygen sensor protein AfGcHK is a globin-coupled histidine kinase in the soil bacterium Anaeromyxobacter sp. Fw109-5. Its C-terminal functional domain exhibits autophosphorylation activity induced by oxygen binding to the heme Fe(II) complex located in the oxygen-sensing N-terminal globin domain. A detailed understanding of the signal transduction mechanisms in heme-containing sensor proteins remains elusive. Here, we investigated the role of the globin domain’s dimerization interface in signal transduction in AfGcHK. We present a crystal structure of a monomeric imidazole-bound AfGcHK globin domain at 1.8 Å resolution, revealing that the helices of the wild-type globin dimer are under tension and suggesting that Tyr-15 plays a role in both this tension and the globin domain’s dimerization. Biophysical experiments revealed that whereas the isolated wild-type globin domain is dimeric in solution, the Y15A and Y15G variants in which Tyr-15 is replaced with Ala or Gly, respectively, are monomeric. Additionally, we found that although the dimerization of the full-length protein is preserved via the kinase domain dimerization interface in all variants, full-length AfGcHK variants bearing the Y15A or Y15G substitutions lack enzymatic activity. The combined structural and biophysical results presented here indicate that Tyr-15 plays a key role in the dimerization of the globin domain of AfGcHK and that globin domain dimerization is essential for internal signal transduction and autophosphorylation in this protein. These findings provide critical insights into the signal transduction mechanism of the histidine kinase AfGcHK from Anaeromyxobacter.

Journal Keywords: dimerization interface; globin; heme-based oxygen sensor; two component system; signal transduction; cell signaling; heme; bacterial protein kinase; histidine kinase; crystal structure

Subject Areas: Biology and Bio-materials


Instruments: I02-Macromolecular Crystallography