The receiver domain of sensor histidine kinase of CKI1: its dynamic structure and binding specificity.
|Year of publication||2010|
|Type||Article in Proceedings|
|MU Faculty or unit|
|Keywords||vícekomponentní fosforylace, 3-D struktura, NMR analýza|
|Description||In Arabidopsis multistep phosphorelay (MSP) signalling, the signal is transferred from sensor histidine kinase (HK) via histidine-containing phosphotransfer proteins (AHP1-5) to nuclear response regulators. In contrast to bacteria, MSP protein interactions in plants are supposed to be rather non-specific. Using both in vivo and in vitro assays we have found that the C-terminal receiver domain of HK CKI1 (CKI1RD) interacts with AHP2, 3 and 5 with different affinities. We determined crystal structure of free CKI1RD and CKI1RD in a complex with Mg2+. We have found that CKI1RD shares a particular similarity with the only known structure of plant HK, ETR1RD, with the main differences in the loop 3. Structural dynamics of CKIRD in solution was studied in detail by NMR in the absence or presence of Mg2+ that was shown to be indispensable to the transphosphorylation activity and BeF3-, the isomorphous stable phosphate analogue. Finally, using in vitro assay (indirect ELISA) we have found, that presence of Mg2+ and acetyl phosphate on one side and BeF3- on the other one affects the interaction of CKI1RD with its downstream signalling partners and may modulate AHP binding specificity in MSP signalling. Observed structural changes, their dynamics and potential influence on signal transduction specificity via MSP in plants will be discussed in detail.|