This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hendriks, J. Articles by Hellingwerf, K. J. Search for Related Content PubMed PubMed Citation Articles by Hendriks, J. Articles by Hellingwerf, K. J.

Deuterium Isotope Effects in the Photocycle Transitions of the Photoactive Yellow Protein

Johnny Hendriks^*, Ivo H. M. van Stokkum and Klaas J. Hellingwerf

^* Department of Biochemistry and Molecular Biology, Faculty of Sciences, Vrije Universiteit, De Boelelaan 1083, 1081 HV Amsterdam, Department of Physics Applied Computer Science, Faculty of Sciences, Vrije Universiteit, De Boelelaan 1081, 1081 HV Amsterdam, and Laboratory for Microbiology, Swammerdam Institute for Life Sciences (SILS), BioCentrum, University of Amsterdam, Nieuwe Achtergracht 166, 1018 WV Amsterdam, The Netherlands

Correspondence: Prof. Dr. Klaas J. Hellingwerf, Laboratory for Microbiology, Nieuwe Achtergracht 166, 1018 WV Amsterdam, The Netherlands. Tel.: +31-20-5257055, Fax: +31-20-5257056; E-mail: K.Hellingwerf{at}science.uva.nl.

The Photoactive Yellow Protein (PYP) from Halorhodospira halophila (formerly Ectothiorhodospira halophila) is increasingly used as a model system. As such, a thorough understanding of the photocycle of PYP is essential. In this study we have combined information from pOH- (or pH-) dependence and (kinetic) deuterium isotope effects to elaborate on existing photocycle models. For several characteristics of PYP we were able to make a distinction between pH- and pOH-dependence, a nontrivial distinction when comparing data from samples dissolved in H₂O and D₂O. It turns out that most characteristics of PYP are pOH-dependent. We confirmed the existence of a pB' intermediate in the pR to pB transition of the photocycle. In addition, we were able to show that the pR to pB' transition is reversible, which explains the previously observed biexponential character of the pR-to-pB photocycle step. Also, the absorption spectrum of pB' is slightly red-shifted with respect to pB. The recovery of the pG state is accompanied by an inverse kinetic deuterium isotope effect. Our interpretation of this is that before the chromophore can be isomerized, it is deprotonated by a hydroxide ion from solution. From this we propose a new photocycle intermediate, pB^deprot, from which pG is recovered and which is in equilibrium with pB. This is supported in our data through the combination of the observed pOH and pH dependence, together with the kinetic deuterium isotope effect.

This article has been cited by other articles:

				D. Hoersch, H. Otto, C. P. Joshi, B. Borucki, M. A. Cusanovich, and M. P. Heyn Role of a Conserved Salt Bridge between the PAS Core and the N-Terminal Domain in the Activation of the Photoreceptor Photoactive Yellow Protein Biophys. J., September 1, 2007; 93(5): 1687 - 1699. [Abstract] [Full Text] [PDF]

				B. Borucki, C. P. Joshi, H. Otto, M. A. Cusanovich, and M. P. Heyn The Transient Accumulation of the Signaling State of Photoactive Yellow Protein Is Controlled by the External pH Biophys. J., October 15, 2006; 91(8): 2991 - 3001. [Abstract] [Full Text] [PDF]

				S. Yeremenko, I. H. M. van Stokkum, K. Moffat, and K. J. Hellingwerf Influence of the Crystalline State on Photoinduced Dynamics of Photoactive Yellow Protein Studied by Ultraviolet-Visible Transient Absorption Spectroscopy Biophys. J., June 1, 2006; 90(11): 4224 - 4235. [Abstract] [Full Text] [PDF]

				N. Shimizu, Y. Imamoto, M. Harigai, H. Kamikubo, Y. Yamazaki, and M. Kataoka pH-dependent Equilibrium between Long Lived Near-UV Intermediates of Photoactive Yellow Protein J. Biol. Chem., February 17, 2006; 281(7): 4318 - 4325. [Abstract] [Full Text] [PDF]

				J. Vreede, W. Crielaard, K. J. Hellingwerf, and P. G. Bolhuis Predicting the Signaling State of Photoactive Yellow Protein Biophys. J., May 1, 2005; 88(5): 3525 - 3535. [Abstract] [Full Text] [PDF]

				M. Unno, M. Kumauchi, N. Hamada, F. Tokunaga, and S. Yamauchi Resonance Raman Evidence for Two Conformations Involved in the L Intermediate of Photoactive Yellow Protein J. Biol. Chem., June 4, 2004; 279(23): 23855 - 23858. [Abstract] [Full Text] [PDF]

				D. Pan, A. Philip, W. D. Hoff, and R. A. Mathies Time-Resolved Resonance Raman Structural Studies of the pB' Intermediate in the Photocycle of Photoactive Yellow Protein Biophys. J., April 1, 2004; 86(4): 2374 - 2382. [Abstract] [Full Text] [PDF]