Demonstration of a sensory rhodopsin in eubacteria.
We report the first sensory rhodopsin observed in the eubacterial domain, a green light-activated photoreceptor in Anabaena (Nostoc) sp. PCC7120, a freshwater cyanobacterium. The gene encoding the membrane opsin protein of 261 residues (26 kDa) and a smaller gene encoding a soluble protein of 125 residues (14 kDa) are under the same promoter in a single operon. The opsin expressed heterologously in Escherichia coli membranes bound all-trans retinal to form a pink pigment (lambda max 543 nm) with a photochemical reaction cycle of 110 ms half-life (pH 6.8, 18 degrees C). Co-expression with the 14 kDa protein increased the rate of the photocycle, indicating physical interaction with the membrane-embedded rhodopsin, which we confirmed in vitro by affinity enrichment chromatography and Biacore interaction. The pigment lacks the proton donor carboxylate residue in helix C conserved in known retinylidene proton pumps and did not exhibit detectable proton ejection activity. We detected retinal binding to the protein in Anabaena membranes by SDS-PAGE and autofluorography of 3H-labelled all-trans retinal of reduced membranes from the organism. We conclude that Anabaena rhodopsin functions as a photosensory receptor in its natural environment, and suggest that the soluble 14 kDa protein transduces a signal from the receptor. Therefore, unlike the archaeal sensory rhodopsins, which transmit signals by transmembrane helix-helix interactions with membrane-embedded transducers, the Anabaena sensory rhodopsin may signal through a soluble cytoplasmic protein, analogous to higher animal visual pigments.
Jung
Kwang-Hwan
KH
Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas-Houston Medical School, Houston, TX 77030, USA.
Trivedi
Vishwa D
VD
Spudich
John L
JL
D000595:Amino Acid Sequence; D017033:Anabaena / Q000235:genetics / Q000378:metabolism*; D001059:Apoproteins / Q000378:metabolism; D001692:Biological Transport; D004926:Escherichia coli / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010777:Photochemistry / Q000379:methods; D011485:Protein Binding / Q000502:physiology; D011522:Protons; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012168:Retinal Pigments / Q000378:metabolism; D012172:Retinaldehyde / Q000378:metabolism; D025603:Sensory Rhodopsins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D012995:Solubility
D001059:Apoproteins; D008565:Membrane Proteins; D011522:Protons; D011994:Recombinant Proteins; D012168:Retinal Pigments; D025603:Sensory Rhodopsins; D012172:Retinaldehyde