title: On the involvement of single-bond rotation in the primary photochemistry of photoactive yellow protein.
abstract: Prior experimental observations, as well as theoretical considerations, have led to the proposal that C(4)-C(7) single-bond rotation may play an important role in the primary photochemistry of photoactive yellow protein (PYP). We therefore synthesized an analog of this protein's 4-hydroxy-cinnamic acid chromophore, (5-hydroxy indan-(1E)-ylidene)acetic acid, in which rotation across the C(4)-C(7) single bond has been locked with an ethane bridge, and we reconstituted the apo form of the wild-type protein and its R52A derivative with this chromophore analog. In PYP reconstituted with the rotation-locked chromophore, 1), absorption spectra of ground and intermediate states are slightly blue-shifted; 2), the quantum yield of photochemistry is ∼60% reduced; 3), the excited-state dynamics of the chromophore are accelerated; and 4), dynamics of the thermal recovery reaction of the protein are accelerated. A significant finding was that the yield of the transient ground-state intermediate in the early phase of the photocycle was considerably higher in the rotation-locked samples than in the corresponding samples reconstituted with p-coumaric acid. In contrast to theoretical predictions, the initial photocycle dynamics of PYP were observed to be not affected by the charge of the amino acid residue at position 52, which was varied by 1), varying the pH of the sample between 5 and 10; and 2), site-directed mutagenesis to construct R52A. These results imply that C(4)-C(7) single-bond rotation in PYP is not an alternative to C(7)=C(8) double-bond rotation, in case the nearby positive charge of R52 is absent, but rather facilitates, presumably with a compensatory movement, the physiological Z/E isomerization of the blue-light-absorbing chromophore.
authors: Department of Physics and Astronomy, Faculty of Sciences, Vrije Universiteit Amsterdam, Amsterdam, The Netherlands.
authors: Andreas D
authors: AD
authors: Stahl
authors: Marijke
authors: M
authors: Hospes
authors: Kushagra
authors: K
authors: Singhal
authors: Ivo
authors: I
authors: van Stokkum
authors: Rienk
authors: R
authors: van Grondelle
authors: Marie Louise
authors: ML
authors: Groot
authors: Klaas J
authors: KJ
authors: Hellingwerf
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D020607:Halorhodospira halophila; D006863:Hydrogen-Ion Concentration; D055668:Photochemical Processes; D020283:Photoreceptors, Microbial / Q000737:chemistry* / Q000378:metabolism; D012399:Rotation; D013057:Spectrum Analysis
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; C050162:photoactive yellow protein, Bacteria