title: Visualizing reaction pathways in photoactive yellow protein from nanoseconds to seconds.
abstract: Determining 3D intermediate structures during the biological action of proteins in real time under ambient conditions is essential for understanding how proteins function. Here we use time-resolved Laue crystallography to extract short-lived intermediate structures and thereby unveil signal transduction in the blue light photoreceptor photoactive yellow protein (PYP) from Halorhodospira halophila. By analyzing a comprehensive set of Laue data during the PYP photocycle (forty-seven time points from one nanosecond to one second), we track all atoms in PYP during its photocycle and directly observe how absorption of a blue light photon by its p-coumaric acid chromophore triggers a reversible photocycle. We identify a complex chemical mechanism characterized by five distinct structural intermediates. Structural changes at the chromophore in the early, red-shifted intermediates are transduced to the exterior of the protein in the late, blue-shifted intermediates through an initial "volume-conserving" isomerization of the chromophore and the progressive disruption of hydrogen bonds between the chromophore and its surrounding binding pocket. These results yield a comprehensive view of the PYP photocycle when seen in the light of previous biophysical studies on the system.
authors.affiliation: Department of Chemistry and School of Molecular Science (BK21), Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701, South Korea. hyotcherl.ihee@kaist.ac.kr
authors.forename: Hyotcherl
authors.initials: H
authors.lastname: Ihee
authors.forename: Sudarshan
authors.initials: S
authors.lastname: Rajagopal
authors.forename: Vukica
authors.initials: V
authors.lastname: Srajer
authors.forename: Reinhard
authors.initials: R
authors.lastname: Pahl
authors.forename: Spencer
authors.initials: S
authors.lastname: Anderson
authors.forename: Marius
authors.initials: M
authors.lastname: Schmidt
authors.forename: Friedrich
authors.initials: F
authors.lastname: Schotte
authors.forename: Philip A
authors.initials: PA
authors.lastname: Anfinrud
authors.forename: Michael
authors.initials: M
authors.lastname: Wulff
authors.forename: Keith
authors.initials: K
authors.lastname: Moffat
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism* / Q000502:physiology; D003373:Coumaric Acids / Q000378:metabolism; D003461:Crystallography; D007536:Isomerism; D007700:Kinetics; D008958:Models, Molecular; D017785:Photons; D020283:Photoreceptors, Microbial / Q000378:metabolism* / Q000502:physiology; D011422:Propionates; D015398:Signal Transduction / Q000502:physiology*; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; D003373:Coumaric Acids; D020283:Photoreceptors, Microbial; D011422:Propionates; C050162:photoactive yellow protein, Bacteria; C495469:p-coumaric acid