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Instruments / Technical Area	Publication Details	Published	Actions
I02-Macromolecular Crystallography	Disruption of the dimerization interface of the sensing domain in the dimeric heme-based oxygen sensor Af GcHK abolishes bacterial signal transduction Tereza Skalova , Alzbeta Lengalova , Jan Dohnalek , Karl Harlos , Peter Mihalcin , Petr Kolenko , Martin Stranava , Jan Blaha , Toru Shimizu , Markéta Martínková Journal Of Biological Chemistry DOI: 10.1074/jbc.RA119.011574 Diamond Proposal Number(s): [10627] Open Access Show Abstract ▼ 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.	Dec 2019

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I02-Macromolecular Crystallography

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

Tereza Skalova , Alzbeta Lengalova , Jan Dohnalek , Karl Harlos , Peter Mihalcin , Petr Kolenko , Martin Stranava , Jan Blaha , Toru Shimizu , Markéta Martínková

Journal Of Biological Chemistry

DOI: 10.1074/jbc.RA119.011574

Diamond Proposal Number(s): [10627]