Proteorhodopsin in living color: diversity of spectral properties within living bacterial cells.
Proteorhodopsin is a family of over 50 proteins that provide phototrophic capability to marine bacteria by acting as light-powered proton pumps. The potential importance of proteorhodopsin to global ocean ecosystems and the possible applications of proteorhodopsin in optical data storage and optical signal processing have spurred diverse research in this new family of proteins. We show that proteorhodopsin expressed in Escherichia coli is functional and properly inserted in the membrane. At high expression levels, it appears to self-associate. We present a method for determining spectral properties of proteorhodopsin in intact E. coli cells that matches results obtained with detergent-solubilized, purified proteins. Using this method, we observe distinctly different spectra for protonated and deprotonated forms of 21 natural proteorhodopsin proteins in intact E. coli cells. Upon protonation, the wavelength maxima red shifts between 13 and 53 nm. We find that pKa values between 7.1 and 8.5 describe the pH-dependent spectral shift for all of the 21 natural variants of proteorhodopsin. The wavelength maxima of the deprotonated forms of the 21 natural proteorhodopsins cluster in two sequence-related groups: blue proteorhodopsins (B-PR) and green proteorhodopsins (G-PR). The site-directed substitution Leu105Gln in Bac31A8 proteorhodopsin shifts this G-PR's wavelength maximum to a wavelength maximum the same as that of the B-PR Hot75m1 proteorhodopsin. The site-directed substitution Gln107Leu in Hot75m1 proteorhodopsin shifts this B-PR's wavelength maximum to a wavelength maximum as that of Bac31A8 proteorhodopsin.
Kelemen
Bradley R
BR
Genencor International, Inc, 925 Page Mill Road, Palo Alto, CA 94304, USA.
Du
Mai
M
Jensen
Rasmus B
RB
D001425:Bacterial Outer Membrane Proteins; D003001:Cloning, Molecular; D004926:Escherichia coli; D014644:Genetic Variation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D018272:Porins; D011991:Receptors, Virus / Q000737:chemistry* / Q000235:genetics; D012243:Rhodopsin / Q000737:chemistry* / Q000235:genetics; D025604:Rhodopsins, Microbial; D013053:Spectrophotometry; D013329:Structure-Activity Relationship
D001425:Bacterial Outer Membrane Proteins; D018272:Porins; D011991:Receptors, Virus; D025604:Rhodopsins, Microbial; C016532:maltoporins; C413548:proteorhodopsin; D012243:Rhodopsin