Positioning proton-donating residues to the Schiff-base accelerates the M-decay of pharaonis phoborhodopsin expressed in Escherichia coli.
Phoborhodopsin (also called sensory rhodopsin II, sR-II) is a receptor for the negative phototaxis of Halobacterium salinarum (pR), and pharaonis phoborhodopsin (ppR) is the corresponding receptor of Natronobacterium pharaonis. pR and ppR are retinoid proteins and have a photocycle similar to that of bacteriorhodopsin (bR). A major difference between the photocycle of the ion pump bR and the sensor pR or ppR is found in their turnover rates which are much faster for bR. A reason for this difference might be found in the lack of a proton-donating residue to the Schiff base which is formed between the lysine of the opsin and retinal. To reconstruct a bR-like photochemical behavior, we expressed ppR mutants in Escherichia coli in which proton-donating groups have been reintroduced into the cytoplasmic proton channel. In measurement of the photocycle it could be shown that the F86D mutant of ppR (Phe86 was substituted by Asp) showed a faster decay of M-intermediate than the wild-type, which was even accelerated in the F86D/L40T double mutant.
Iwamoto
M
M
Laboratory of Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Shimono
K
K
Sumi
M
M
Kamo
N
N
D019843:Archaeal Proteins; D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002338:Carotenoids; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D019617:Halobacterium salinarum / Q000378:metabolism; D025602:Halorhodopsins; D007700:Kinetics; D019633:Natronobacterium / Q000235:genetics / Q000378:metabolism; D010777:Photochemistry; D017354:Point Mutation; D011522:Protons; D012545:Schiff Bases / Q000737:chemistry / Q000378:metabolism; D025603:Sensory Rhodopsins
D019843:Archaeal Proteins; D017931:DNA Primers; D025602:Halorhodopsins; D011522:Protons; D012545:Schiff Bases; D025603:Sensory Rhodopsins; C061508:sensory rhodopsin II protein, archaeal; D002338:Carotenoids; D001436:Bacteriorhodopsins