Interaction of the Halobacterial Transducer to a Halorhodopsin Mutant Engineered so as to Bind the Transducer : Cl- Circulation within the Extracellular Channel

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An alkali-halophilic archaeum, Natronomonas pharaonis, contains two rhodopsins that are halorhodopsin (phR), a light-driven inward Cl- pump and phoborhodopsin (ppR), the receptor of negative phototaxis functioning by forming a signaling complexes with a transducer, pHtrII. Previously, we reported that the phR double mutant, P240T/F250YphR, can bind with pHtrII [Y. Sudo et al. (2006) J. Mol. Biol. 357, 1274-1282]. This mutant itself can transport Cl-, while the net transport was stopped upon formation of the complex. The flash photolysis data were analyzed by a scheme in which phR ⇒ P1 → P2 → P3 → P4 → phR. The P3 of the wild-type and the double mutant contained two components, X- and O-intermediates. After the complex formation, however, the P3 of the double mutant lacked the X-intermediate. These observations imply that the X-intermediate (probably the N-intermediate) is the state having Cl- in the cytoplasmic binding site and that the complex undergoes an extracellular Cl- circulation due to the inhibition of formation of the X-intermediate.