title: Measurement and global analysis of the absorbance changes in the photocycle of the photoactive yellow protein from Ectothiorhodospira halophila.
abstract: The photocycle of the photoactive yellow protein (PYP) from Ectothiorhodospira halophila was examined by time-resolved difference absorption spectroscopy in the wavelength range of 300-600 nm. Both time-gated spectra and single wavelength traces were measured. Global analysis of the data established that in the time domain between 5 ns and 2 s only two intermediates are involved in the room temperature photocycle of PYP, as has been proposed before (Meyer T.E., E. Yakali, M. A. Cusanovich, and G. Tollin. 1987. Biochemistry. 26:418-423; Meyer, T. E., G. Tollin, T. P. Causgrove, P. Cheng, and R. E. Blankenship. 1991. Biophys. J. 59:988-991). The first, red-shifted intermediate decays biexponentially (60% with tau = 0.25 ms and 40% with tau = 1.2 ms) to a blue-shifted intermediate. The last step of the photocycle is the biexponential (93% with tau = 0.15 s and 7% with tau = 2.0 s) recovery to the ground state of the protein. Reconstruction of the absolute spectra of these photointermediates yielded absorbance maxima of about 465 and 355 nm for the red- and blue-shifted intermediate with an epsilon max at about 50% and 40% relative to the epsilon max of the ground state. The quantitative analysis of the photocycle in PYP described here paves the way to a detailed biophysical analysis of the processes occurring in this photoreceptor molecule.
authors: Department of Microbiology, E.C. Slater Institute, BioCentrum Amsterdam, University of Amsterdam, The Netherlands.
authors: W D
authors: WD
authors: Hoff
authors: I H
authors: IH
authors: van Stokkum
authors: H J
authors: HJ
authors: van Ramesdonk
authors: M E
authors: ME
authors: van Brederode
authors: A M
authors: AM
authors: Brouwer
authors: J C
authors: JC
authors: Fitch
authors: T E
authors: TE
authors: Meyer
authors: R
authors: R
authors: van Grondelle
authors: K J
authors: KJ
authors: Hellingwerf
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D002841:Chromatiaceae / Q000378:metabolism*; D003624:Darkness; D007700:Kinetics; D008027:Light; D008433:Mathematics; D008962:Models, Theoretical; D020283:Photoreceptors, Microbial; D013053:Spectrophotometry; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; C050162:photoactive yellow protein, Bacteria