Biochemical characterisation of the cyanobacterial Hik2-Rre1 two-component regulatory system.
MetadataShow full item record
Two-component signal transduction systems (TCS) consist of a sensor histidine kinase and a response regulator. TCS are ubiquitous in prokaryotes, but found only in some eukaryotes. TCS mediate adaptation to various environmental changes in bacteria, plants, fungi, and protists. Histidine kinase 2 (Hik2) is a sensor histidine kinase found in all cyanobacteria. The Hik2 homologue known as Chloroplast Sensor Kinase is found in algae and plants, where it is encoded by the nuclear genome and it is targeted to chloroplasts. CSK couples the redox state of the photosynthetic electron transport chain to chloroplast gene transcription. This thesis describes biochemical characterisation of the signal transduction mechanism of Hik2 and its response regulator (Rre) partners in order to clarify the Hik2-Rre two-component signal transduction pathway. Results presented in this thesis illustrate that the autophosphorylation activity of the full-length Hik2 protein is specifically inhibited by sodium ions. An autophosphorylation event of a histidine kinase is the result of homodimerisation and is followed by trans or cis-autophosphorylation of each monomer on its conserved histidine residue. Chemical crosslinking revealed that the Hik2 protein exists predominantly as a phosphorylated (autokinase active) monomer, tetramer, and higher-order oligomeric complexes. The functions of these different oligomeric states of Hik2 are also discussed. From a previous study, which was based on an observation from a yeast two-hybrid assay, Hik2 was proposed to form a two-component pair with Rre1 and RppA. However, no further evidence was presented to support either direct interaction or direct phosphotransfer activity of the Hik2-Rre pair. This thesis confirms interaction of Hik2-Rre1 and Hik2-RppA two-component ! %! pairs using an in vitro pull-down assay and phosphotransfer kinetics. Finally, a model is proposed for the Hik2 based two-component signal transduction pathway.
AuthorsIbrahim, Iskander Mohamed
- Theses