Article,
A non-hydrolyzable ATP derivative generates a stable complex in a light-inducible two component system
J Biol Chem, (October 2009)
DOI: 10.1074/jbc.M109.017772
Abstract
Isothermal calorimetry (ITC) measurements yielded the binding constants during complex formation of light-inducible histidine kinases (HK) and their cognate CheY-type response regulators (RR). HK-RR interactions represent the core function of the bacterial two component system, which is also present in many bacterial phytochromes. Here, we have studied the recombinant forms of phytochromes CphA and CphB from the cyanobacterium Tolypothrix PCC7601 and their cognate RRs RcpA and RcpB. Interaction between the two reaction partners (HK and RR) was studied in presence and absence of ATP. A complex formation was observable in the presence of ATP, but specific interactions were only found, when a non-hydrolysable ATP-derivative was added to the mixture. Also, the incubation of the HK domain alone (expressed as a recombinant protein) with the RR did not yield specific interactions, indicating that the HK domain is only active as component of the full-length phytochrome. Considering also previous studies on the same proteins (Hubschmann et al., 2001, Eur. J. Biochem. 268, 3383-3389) we now conclude that the HK domains of these phytochromes are active only when the chromophore domain is in its Pr form. The formerly documented phosphate transfer between the HK-domain and the RR takes place via an only transient protein-protein complex, which becomes detectable by ITC in the presence of non-hydrolysable ATP derivative. This finding is of interest also in relation to the function of some (blue light-sensitive) photoreceptors that carry the HK domain and the RR fused together in one single protein.
