title: Functional expression and site-directed mutagenesis of photoactive yellow protein.
abstract: The gene encoding photoactive yellow protein (PYP) was isolated from Ectothiorhodospira halophila, and a high-level expression system for PYP was constructed in Escherichia coli. The molecular weight and the absorption spectrum of PYP expressed in E. coli were identical with those of the native PYP isolated from E. halophila. The amino acid residues which might interact with the chromophore (Tyr42, Glu46, Thr50, Arg52, and Cys69) were mutated by site-directed mutagenesis and the absorption spectra of these mutants were examined to study the chromophore/protein interaction in PYP. The former three substitutions (Y42F, E46Q, and T50V) brought about red-shifts of the absorption spectra, but the substitution of Arg52 (R52Q) brought about no change and that of Cys69 (C69S) led to no formation of pigments. These results suggest that Tyr42, Glu46, and Thr50 strongly interact with the chromophore, while Arg52 does not contribute the color tuning of PYP.
authors.affiliation: Department of Earth and Space Science, Graduate School of Science, Osaka University, Toyonaka.
authors.forename: K
authors.initials: K
authors.lastname: Mihara
authors.forename: O
authors.initials: O
authors.lastname: Hisatomi
authors.forename: Y
authors.initials: Y
authors.lastname: Imamoto
authors.forename: M
authors.initials: M
authors.lastname: Kataoka
authors.forename: F
authors.initials: F
authors.lastname: Tokunaga
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D002841:Chromatiaceae / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed / Q000235:genetics*; D020283:Photoreceptors, Microbial; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D012995:Solubility; D013053:Spectrophotometry
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; D011994:Recombinant Proteins; C050162:photoactive yellow protein, Bacteria