Subproject B05: H2O2 perception and signal transduction: The specific role of AHK5
Principal investigators:
Prof. Dr. Klaus Harter
Universität Tübingen
ZMBP, Pflanzenphysiologie
Auf der Morgenstelle 1,
72076 Tübingen
Tel 07071 - 29 72605
Fax 07071 - 29 3287
klaus.harter@zmbp.uni-tuebingen.de
Prof. Dr. Thilo Stehle
Universität Tübingen
IFIB, Strukturbiologie
Hoppe-Seyler-Str. 4, 72076 Tübingen
Tel 07071 - 29 73043
FAX 07071 - 29 5565
Summary:
Plant histidine kinases (HKs) are multifunctional proteins that sense specific stimuli and generate a specific signal output via the Two-Component System (TCS) network. HKs are typically homodimers that possess His autophosphorylation and His-to-Asp phosphorelay activities. Despite their critical functions, plant HKs are poorly understood at the structural and functional level with respect to how they sense their specific stimuli and how they convert the sensed information into specific intracellular signalling.
In the previous funding period, we could extend our knowledge on the involvement of AHK5 in H2O2-dependent physiological and developmental processes. Furthermore, we were able show that Cysteine 3 (Cys3) is required for AHK5 His kinase activity in vivo and H2O2-dependent disulphide bridge formation of the N-terminal AHK5 domain in vitro. In combination with additional biochemical, cell biological and protein-protein interaction data, this suggests that AHK5 is likely to function as a H2O2-/redox-regulated HK.
In the upcoming funding period, we will solve structures of AHK5 domains, and we will combine the structural information with functional studies. We will also initiate structural studies of full-length AHK5 expressed in insect cells. These experiments should allow us to elucidate how AHK5 senses H2O2 and how the protein measures the redox state in vitro and within the cell. Furthermore, the structural data will suggest site-directed mutagenesis experiments that will help to establish to which extent AHK5 His kinase activity, phosphorelay, and cross-talk to classical Ser/Thr/Tyr phosphorylation cascades are required for the AHK5-specific physiological and developmental outputs. Furthermore, it is planned to identify other proteins that contribute to AHK5-specific signalling and characterize their function in the context of H2O2 as signalling molecule.