title: Protein kinetics: structures of intermediates and reaction mechanism from time-resolved x-ray data.
abstract: We determine the number of authentic reaction intermediates in the later stages of the photocycle of photoactive yellow protein at room temperature, their atomic structures, and a consistent set of chemical kinetic mechanisms, by analysis of a set of time-dependent difference electron density maps spanning the time range from 5 micros to 100 ms. The successful fit of exponentials to right singular vectors derived from a singular value decomposition of the difference maps demonstrates that a chemical kinetic mechanism holds and that structurally distinct intermediates exist. We identify two time-independent difference maps, from which we refine the structures of the corresponding intermediates. We thus demonstrate how structures associated with intermediate states can be extracted from the experimental, time-dependent crystallographic data. Stoichiometric and structural constraints allow the exclusion of one kinetic mechanism proposed for the photocycle but retain other plausible candidate kinetic mechanisms.
authors.affiliation: Physikdepartment E17, Technische Universität München, 85747 Garching, Germany. marius@hexa.e17.physik.tu-muenchen.de
authors.forename: Marius
authors.initials: M
authors.lastname: Schmidt
authors.forename: Reinhard
authors.initials: R
authors.lastname: Pahl
authors.forename: Vukica
authors.initials: V
authors.lastname: Srajer
authors.forename: Spencer
authors.initials: S
authors.lastname: Anderson
authors.forename: Zhong
authors.initials: Z
authors.lastname: Ren
authors.forename: Hyotcherl
authors.initials: H
authors.lastname: Ihee
authors.forename: Sudarshan
authors.initials: S
authors.lastname: Rajagopal
authors.forename: Keith
authors.initials: K
authors.lastname: Moffat
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D018360:Crystallography, X-Ray; D007700:Kinetics; D020283:Photoreceptors, Microbial / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; C050162:photoactive yellow protein, Bacteria