title: L105K mutant of proteorhodopsin.
abstract: Proteorhodopsin (PR) is a light-driven proton pump found in marine bacteria. Thousands of PRs are classified into blue-absorbing (λ(max) ~ 490 nm) and green-absorbing (λ(max) ~ 525 nm) PR, and the color determinant is known to be at position 105, where blue-absorbing and green-absorbing PR possess Gln and Leu, respectively. Position 105 is in contact with the retinal chromophore in the hydrophobic region of the cytoplasmic side. In this paper, we have introduced a positively charged lysine group at position 105, which is the first report of the introduction of a positively charged group into the hydrophobic cytoplasmic domain in microbial rhodopsins. The L105K mutant PR shows an ~21 nm red shift (λ(max) ~ 549 nm) at pH 7.0, and the pK(a) of the counterion (7.2) does not change significantly compared to that of wild-type PR (6.8). The analysis of thermal stability shows that the mutation causes some destabilization of structure, but the mutant is more stable toward hydroxylamine reaction than the wild type. The flash photolysis measurement at pH 9.0 shows that the decay of the M intermediate of L105K is ~3 times slower than that of the wild type. The slow M decay possibly originates from the perturbation of the proton donor (Glu108) and the retinal Schiff base due to positioning of a positively charged lysine group in the proton transfer pathway. The perturbation of proton transport is also observed when we measure light-induced proton pumping. The rate of proton transport in L105K mutant is 6 times slower than that of the wild type, which corroborates our flash photolysis result.
authors.affiliation: Department of Frontier Materials, Nagoya Institute of Technology, Showa-ku, Nagoya 466-8555, Japan.
authors.forename: Tushar Kanti
authors.initials: TK
authors.lastname: Maiti
authors.forename: Keisuke
authors.initials: K
authors.lastname: Yamada
authors.forename: Keiichi
authors.initials: K
authors.lastname: Inoue
authors.forename: Hideki
authors.initials: H
authors.lastname: Kandori
mesh_terms: D005973:Glutamine / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007930:Leucine / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D012243:Rhodopsin / Q000737:chemistry* / Q000235:genetics; D025604:Rhodopsins, Microbial; D012545:Schiff Bases / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D025604:Rhodopsins, Microbial; D012545:Schiff Bases; C413548:proteorhodopsin; D005973:Glutamine; D012243:Rhodopsin; D007930:Leucine