Histidine kinases are the bacterial counterpart to eukaryotic Ser/Thr and Tyr protein kinases, all of which function as signal transducers required for cellular response and adaptation to external signals (stresses). Typically bacterial histidine kinases (excluding a histidine kinase involved in chemotaxis) are transmembrane signal transducers that regulate the phosphorylation state of their own cognate response regulators, which function mostly as transcription factors. This in turn regulates (activates or represses) a specific gene required for adaptation to the stresses. In contrast to Ser/Thr and Tyr protein kinases, a high-energy phosphoryl group is relayed from ATP through a histidine kinase to a response regulator during signal transduction. Therefore, the histidine kinase-response regulator systems are referred as """"""""the two-component His-Asp phosphorelay signal transduction system"""""""". Of these, the EnvZ-OmpR system from Escherichia coli is one of the most extensively investigated systems. EnvZ is an osmo-sensing histidine kinase and the NMR three-dimensional structures of its cytoplasmic kinase domains have been determined.
In Aim 1, we will continue the structural studies on the periplasmic receptor domain and the cytoplasmic linker domain to comprehend the global three-dimensional structure of the EnvZ dimer in the membrane. We will also construct hybrid signal transducers between EnvZ and Tar, a chemotaxis Asp sensor to obtain mechanistic insights into the precise molecular mechanism of regulation of the histidine kinase activity by a signal input.
In Aim 2, we will decipher the precise mode of binding of phosphorylated OmpR (OmpR-P) to six OmpR-P binding sites in the ompF promoter to prove a model called """"""""galloping model"""""""" predicted from the preliminary results. We will also analyze the reciprocal regulatory mechanism for the ompF and ompC outer membrane porin genes at the level of transcription by OmpR-P and at the level of translation by two antisense RNAs, micF and micC RNAs by RT-PCR. ? ?
| Yoshida, Takeshi; Phadtare, Sangita; Inouye, Masayori (2007) The design and development of Tar-EnvZ chimeric receptors. Methods Enzymol 423:166-83 |
| Yoshida, Takeshi; Phadtare, Sangita; Inouye, Masayori (2007) Functional and structural characterization of EnvZ, an osmosensing histidine kinase of E. coli. Methods Enzymol 423:184-202 |
| Yoshida, Takeshi; Qin, Ling; Egger, Linda A et al. (2006) Transcription regulation of ompF and ompC by a single transcription factor, OmpR. J Biol Chem 281:17114-23 |
