Structural characterization of the heme‐based oxygen sensor, AfGcHK, its interactions with the cognate response regulator, and their combined mechanism of action in a bacterial two‐component signaling system. Issue 10 (23rd June 2016)

• Record Type: Journal Article
• Authors: Stranava, Martin; Martínek, Václav; Man, Petr; Fojtikova, Veronika; Kavan, Daniel; Vaněk, Ondřej; Shimizu, Toru; Martinkova, Marketa
• Is Part Of: Proteins. Volume 84:Issue 10(2016)
• Page Start: 1375
• Page End: 1389
• Publication Date: 2016-06-23
• Subjects: heme‐based oxygen sensor -- histidine kinase -- two‐component signal transduction system -- protein–protein interaction -- hydrogen‐deuterium exchange -- analytical ultracentrifugation -- homology modeling -- protein–protein docking; Proteins -- Periodicals; Proteins -- Periodicals; 572.6
• DOI: 10.1002/prot.25083
• Languages: English
• ISSNs: 0887-3585

Abstract

ABSTRACT: The oxygen sensor histidine kinase Af GcHK from the bacterium Anaeromyxobacter sp. Fw 109‐5 forms a two‐component signal transduction system together with its cognate response regulator (RR). The binding of oxygen to the heme iron of its N‐terminal sensor domain causes the C‐terminal kinase domain of Af GcHK to autophosphorylate at His183 and then transfer this phosphate to Asp52 or Asp169 of the RR protein. Analytical ultracentrifugation revealed that Af GcHK and the RR protein form a complex with 2:1 stoichiometry. Hydrogen‐deuterium exchange coupled to mass spectrometry (HDX‐MS) suggested that the most flexible part of the whole Af GcHK protein is a loop that connects the two domains and that the heme distal side of Af GcHK, which is responsible for oxygen binding, is the only flexible part of the sensor domain. HDX‐MS studies on the Af GcHK:RR complex also showed that the N‐side of the H9 helix in the dimerization domain of the Af GcHK kinase domain interacts with the helix H1 and the β‐strand B2 area of the RR protein's Rec1 domain, and that the C‐side of the H8 helix region in the dimerization domain of the Af GcHK protein interacts mostly with the helix H5 and β‐strand B6 area of the Rec1 domain. The Rec1 domain containing the phosphorylable Asp52 of the RR protein probably has a significantly higher affinity for Af GcHK than the Rec2 domain. We speculate that phosphorylation at Asp52 changes the overall structure of RR such that the Rec2 area containing the second phosphorylation site (Asp169) can also interact with Af GcHK. Proteins 2016; 84:1375–1389. © 2016 Wiley Periodicals, Inc.