title: pH dependence of the photoactive yellow protein photocycle investigated by time-resolved crystallography.
abstract: Visualizing the three-dimensional structures of a protein during its biological activity is key to understanding its mechanism. In general, protein structure and function are pH-dependent. Changing the pH provides new insights into the mechanisms that are involved in protein activity. Photoactive yellow protein (PYP) is a signaling protein that serves as an ideal model for time-dependent studies on light-activated proteins. Its photocycle is studied extensively under different pH conditions. However, the structures of the intermediates remain unknown until time-resolved crystallography is employed. With the newest beamline developments, a comprehensive time series of Laue data can now be collected from a single protein crystal. This allows us to vary the pH. Here we present the first structure, to our knowledge, of a short-lived protein-inhibitor complex formed in the pB state of the PYP photocycle at pH 4. A water molecule that is transiently stabilized in the chromophore active site prevents the relaxation of the chromophore back to the trans configuration. As a result, the dark-state recovery is slowed down dramatically. At pH 9, PYP stops cycling through the pB state altogether. The electrostatic environment in the chromophore-binding site is the likely reason for this altered kinetics at different pH values.
authors.affiliation: University of Wisconsin-Milwaukee, Milwaukee, Wisconsin, USA.
authors.forename: Shailesh
authors.initials: S
authors.lastname: Tripathi
authors.forename: Vukica
authors.initials: V
authors.lastname: Srajer
authors.forename: Namrta
authors.initials: N
authors.lastname: Purwar
authors.forename: Robert
authors.initials: R
authors.lastname: Henning
authors.forename: Marius
authors.initials: M
authors.lastname: Schmidt
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D018360:Crystallography, X-Ray; D004583:Electrons; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008027:Light; D020283:Photoreceptors, Microbial / Q000737:chemistry* / Q000378:metabolism*; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; C050162:photoactive yellow protein, Bacteria