(Invited) Molecular Mechanisms of Signal Transduction in Heme-Containing Oxygen Sensor Proteins

Abstract

In heme-based gas sensor proteins, heme acts as the sensing site for binding of gaseous molecules, including O2, NO and CO, and indirectly regulates many physiological functions, including the activities of protein kinases, guanylate cyclase, phosphodiesterase, and transcriptional regulatory factors, in response to gas availability. Conceptually, these proteins are always composed of at least two domains: one is a sensor domain (heme-based gas sensing) and the other is a functional domain. However, the structure-function relationship and mechanisms of communication between these domains have not been fully understood. Therefore, we selected a model system, namely a globin-coupled histidine kinase, AfGcHK, in order to study the signal transduction in heme-containing oxygen sensor proteins. The AfGcHK is a part of the two-component signal transduction system from the soil bacterium Anaeromyxobacter sp. Fw109-5. Once the oxygen molecule (as the first signal) binds to the heme iron complex in the sensor domain of AfGcHK, the functional domain is stimulated, leading to autophosphorylation at a conserved His residue in the functional domain. The phosphate group of phosphorylated AfGcHK is then transferred to the cognate response regulator. Several biochemical approaches were utilized in order to study the signal transduction between the sensor and function domains of AfGcHK: (i) enzyme kinetic study, (ii) hydrogen/deuterium exchange experiments associated with mass spectrometry, (iii) cross-linking studies and (iv) small-angle X-ray scattering technique. Overall our results indicated that the coordination and oxidation state of the sensor domain heme iron profoundly affect the enzyme’s catalytic activity of the function domain because they modulate its ATP binding affinity and thus change its k cat/K m ATP value. The possible contact area between sensor and function domains was reveled by hydrogen/deuterium exchange experiments associated with mass spectrometry and cross-linking studies. Small-angle X-ray scattering results offered low-resolution model of the particular domain orientation in solution. All these data together will be discussed in order to illustrate the mechanism of signal transduction in the globin-coupled histidine kinase as a representative of heme-containing oxygen sensor proteins. References: Kitanishi K., Kobayashi K., Uchida T., Ishimori K., Igarashi J., Shimizu T.: Identification and functional and spectral characterization of a globin-coupled histidine kinase from Anaeromyxobacter sp. Fw109-5, J. Biol. Chem . 286, 3522-34 (2011). Martinkova M., Kitanishi K., and Shimizu T.: Heme-Based Globin-Coupled Oxygen Sensors: Linking Oxygen Binding to Functional Regulation of Diguanylate Cyclase, Histidine Kinase and Methyl-accepting Chemotaxis, J. Biol. Chem. 288, 27702–27711 (2013). Fojtikova V., Stranava M., Vos M. H., Liebl U., Hraníček J., Kitanishi K., Shimizu T., Martinkova M.: Kinetic Analysis of a Globin-coupled Histidine Kinase, AfGcHK: Effects of the Heme Iron Complex, Response Regulator and Metal Cations on Autophosphorylation Activity, Biochemistry 54, 5017–5029 (2015). Shimizu T., Yan F., Huang D., Stráňava M., Bartošová M., Fojtíková V., Martínková M.: Molecular Characteristics of Heme-based Gas (O2, NO and CO) Sensors, Chem. Rev. 115, 6491−6533 (2015). Supported in part by Charles University in Prague (UNCE 204025/2012), the Grant Agency of the Czech Republic (grant 15-19883S) and the Grant Agency of Charles University in Prague (grants 756214 and 362115).