A light-driven proton pump from Haloterrigena turkmenica: functional expression in Escherichia coli membrane and coupling with a H+ co-transporter.
A gene encoding putative retinal protein was cloned from Haloterrigena turkmenica (JCM9743). The deduced amino acid sequence was most closely related to that of deltarhodopsin, which functions as a light-driven H+ pump and was identified in a novel strain Haloterrigena sp. arg-4 (K. Ihara, T. Uemura, I. Katagiri, T. Kitajima-Ihara, Y. Sugiyama, Y. Kimura, Y. Mukohata, Evolution of the archaeal rhodopsins: Evolution rate changes by gene duplication and functional differentiation, J. Mol. Biol. 285 (1999) 163-174. GenBank Accession No. AB009620). Thus, we called the present protein H. turkmenica deltarhodopsin (HtdR) in this report. Differing from the Halobacterium salinarum bacteriorhodopsin (bR), functional expression of HtdR was achieved in Escherichia coli membrane with a high yield of 10-15 mg protein/L culture. The photocycle of purified HtdR was similar to that of bR. The photo-induced electrogenic proton pumping activity of HtdR was verified. We co-expressed both HtdR and EmrE, a proton-coupled multi-drug efflux transporter in E. coli, and the cells successfully extruded ethidium, a substrate of EmrE, on illumination.
Kamo
Naoki
N
Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan. nkamo@pharm.hokudai.ac.jp
Hashiba
Tsuyoshi
T
Kikukawa
Takashi
T
Araiso
Tsunehisa
T
Ihara
Kunio
K
Nara
Toshifumi
T
D017920:Antiporters / Q000378:metabolism; D001105:Archaea / Q000378:metabolism; D019843:Archaeal Proteins / Q000737:chemistry; D001436:Bacteriorhodopsins / Q000737:chemistry; D001692:Biological Transport; D002412:Cations; D018432:Drug Resistance, Multiple; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D004996:Ethidium / Q000378:metabolism; D019143:Evolution, Molecular; D020440:Gene Duplication; D006216:Halobacteriaceae / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008027:Light; D008565:Membrane Proteins / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017494:Proton Pumps; D011522:Protons; D012243:Rhodopsin / Q000378:metabolism; D013053:Spectrophotometry; D013997:Time Factors
D017920:Antiporters; D019843:Archaeal Proteins; D002412:Cations; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D017494:Proton Pumps; D011522:Protons; C075436:EmrE protein, E coli; D001436:Bacteriorhodopsins; D012243:Rhodopsin; D004996:Ethidium