Thiol ester-linked p-coumaric acid as a new photoactive prosthetic group in a protein with rhodopsin-like photochemistry

Abstract

Dux, P.; Hard, K.; Devreese, B.; Nugteren-Roodzand, I.M.; Crielaard, W.; Boelens, R.; Beeumen, J.; Kaptein, R.; Hellingwerf, K.J. Published in: Biochemistry DOI:10.1021/bi00251a001Link to publication Citation for published version (APA):Hoff, W. D., Dux, P., Hard, K., Devreese, B., Nugteren-Roodzand, I. M., Crielaard, W., ... Hellingwerf, K. J. (1994). p-Coumaric acid, a new photoactive chromophore of a yellow photoreceptor protein with rhodopsin-like characteristics. Biochemistry, 33, 13959-13963. DOI: 10.1021/bi00251a001 General rightsIt is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons). Disclaimer/Complaints regulationsIf you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: http://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible. would make it the first eubacterial rhodopsin. Here we report the chemical structure of this chromophoric group to be p-coumaric acid, which is covalently bound to a unique cysteine in the apoprotein via a thiol ester bond, and thus not retinal. This makes PYP the first example of a protein containing p-coumaric acid, a metabolite previously found only in plants, as a prosthetic group and establishes the photoactive yellow proteins as a new type of photochemically active receptor molecule.The photoactive yellow proteins (PYP) constitute a responsible for the yellow color of the protein have been advanced (Meyer, 1985;McRee at al, 1989;Van Beeumen et al., 1993), but the true nature of this chromophore et al, 1993) and crystal structure ( M~R~~ et al., 1989) of pyp at 2.4-ij resolution have been and show that the protein is composed of two perpendicular plates of P-sheet, forming a p-clam structure very similar to the fold homologous group of proteins found in many Eubacteria (Meyer, 1985; M e w et al., 1990;Hoff et al., 1994a). The isolated from Ecfothiorhodospira halophila have been studied in some detail. Since PYP was isolated in 1985, a number of proposals concerning the chemical structure of the cofactor structural and photochemical characteristics Of the PYP remained unclear. The amino acid sequence (Van Beeumen

Chemical Reactivity and Spectroscopy of the Thiol Ester-Linked p-Coumaric Acid Chromophore in the Photoactive Yellow Protein from Ectothiorhodospira halophila

“…By absorbance of a photon, PYP is excited into a photocycle that involves a red-shifted and a blue-shifted intermediate (Meyer et al, 1987;Hoff et al, 1994c), very similar to what has been described for the sensory rhodopsins (Spudich & Bogomolni, 1988). However, we have recently determined the PYP cofactor to be a p-coumaric acid molecule [Hoff et al, 1994a;confirmed in Baca et al (1994)] linked to Cys 69 [see Van Beeumen et al (1993)], presumably via a thiol ester bond. Therefore, it is very different from the Schiff base-bound retinal molecule present in the rhodopsins.…”

“…This experiment was repeated using 11 mg of pure PYP. NMR measurements of the resulting chromophore-containing fraction in D 2 O indicated that in this procedure the bond between the chromophore and the apoprotein was broken [ Table 1; see Hoff et al (1994a)], explaining the difference in absorption spectra between this fraction and the pepsin-generated chromopeptide.…”

“…This process is reversible, since reneutralization of the sample leads to the re-formation of the native absorbance band at 446 nm (data not shown). However, this peak is somewhat reduced in intensity, indicating that in addition to the reversible 446 to 410 nm Hoff et al (1994a)]. f Minor amounts.…”

Evidence for trans‐cis isomerization of the p‐coumaric acid chromophore as the photochemical basis of the photocycle of photoactive yellow protein

“…In the ground state pG, the chromophore of PYP is in the trans configuration (referring to the vinyl protons) and in the deprotonated state (the p-hydroxy group), as was indicated by 1H NMR [1] and resonance Raman spectroscopy [12], respectively. It has been proposed that also in the case of PYP lightinduced chromophore photo-isomerization occurs [1,28].…”

“…Photoactive yellow protein (PYP) from the purple sulfur bacterium Ectothiorhodospira halophila is the first eubacteriai photoreceptor to be characterized in detail and has recently been shown to contain a new chromophoric group: thiol ester linked p-coumaric acid [1,2]. This is t he first report of a physiological role for p-coumaric acid in rrokaryotes, a compound previously identified in higher plants, where it plays a central.…”