Interaction of the halobacterial transducer to a halorhodopsin mutant engineered so as to bind the transducer: Cl- circulation within the extracellular channel.
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 complex with a transducer, pHtrII (Sudo Y. et al., J. Mol. Biol. 357 [2006] 1274). Previously, we reported that the phR double mutant, P240T/F250Y(phR), can bind with pHtrII. 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 --> 4 P1 --> P2 --> 4 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 because of the inhibition of formation of the X-intermediate.
Hasegawa
Chisa
C
Laboratory of Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Kikukawa
Takashi
T
Miyauchi
Seiji
S
Seki
Akiteru
A
Sudo
Yuki
Y
Kubo
Megumi
M
Demura
Makoto
M
Kamo
Naoki
N
D000818:Animals; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D002338:Carotenoids / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D005260:Female; D006216:Halobacteriaceae / Q000235:genetics / Q000378:metabolism; D025602:Halorhodopsins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D066298:In Vitro Techniques; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D018462:Photobiology; D010782:Photolysis; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014981:Xenopus
D019843:Archaeal Proteins; D002712:Chlorides; D025602:Halorhodopsins; C434451:HtrII protein, Natronobacterium pharaonis; D011994:Recombinant Proteins; C434450:phototaxis receptor sensory rhodopsin II, Natronobacterium pharaonis; D002338:Carotenoids