Transient proton uptake and release is associated with the photocycle of the photoactive yellow protein from the purple phototrophic bacterium Ectothiorhodospira halophila.
Upon excitation by a laser flash at 445 nm, the photoactive yellow protein (PYP) undergoes a bleach and red-shift occurring in less than 10 ns, undergoes a further bleach in approximately 200 microseconds, and then recolors in approximately 200 ms. A conformational change occurs during photobleaching which exposes a hydrophobic site. We have now shown that this photocycle also involves a net uptake of one proton during formation of the fully bleached second intermediate, followed by an equivalent proton release upon return of PYP to the ground state. Proton uptake lags slightly behind PYP bleaching and is first-order, indicating that the protein conformational change occurs in two steps. The results suggest that a basic residue which is normally buried in the protein interior is transiently exposed to solvent during the PYP photocycle and, as a consequence, undergoes a change in pK. On the basis of the crystal structure of PYP, we propose that this basic residue is lysine 111.
Meyer
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TE
Department of Biochemistry, University of Arizona, Tucson 85721.
Cusanovich
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MA
Tollin
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G
D001426:Bacterial Proteins / Q000378:metabolism* / Q000528:radiation effects; D001962:Bromcresol Purple; D002841:Chromatiaceae / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007834:Lasers; D008027:Light; D020283:Photoreceptors, Microbial; D013053:Spectrophotometry; D013997:Time Factors
D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; C050162:photoactive yellow protein, Bacteria; D001962:Bromcresol Purple