Fakultät für Chemie und Pharmazie
Institut für Physikalische und Theoretische Chemie
Lehrstuhl für Physikalische Chemie

Molecular Spectroscopy and Photochemistry of Sensory Photoreceptors

Prof. Dr. Bernhard Dick
Group Members:
Dr. Uwe Kensy
Dr. Roger-Jan Kutta
Former Group Members:
Dr. Tilman Kottke
Dr. Gilbert Nöll
Dr. Huimin Guo
Cover Paper
DFG-Graduiertenkolleg GRK 640
Sensory Photoreceptors
DFG-Forschergruppe FOR 526
Blue-Light Photoreceptors


The photocycles of biological blue-light photoreceptors.

Living organisms employ photoreactive proteins for two main purposes: energy storage, and sensory functions. Presently, six families of biological photosensors are known. [1] Three of them, the rhodopsins, phytochromes, and xanthopsins, make use of a cis/trans isomerization of the photoreactive cofactor (retinal, tetrapyrrole, or p-coumaric acid). Since 1997, three further classes of photoreceptor protein domains have been found. They all contain a flavin cofactor as chromophore, which makes them sensitive for blue light only. The cryptochromes (CRY) [2,3] are frequently associated with circadian rhythms and contain flavin adenine dinucleotide (FAD), as do the BLUF (blue light sensing using flavin) proteins. [4] The third class are the phototropins, [5] which are composed of (usually two) LOV (light oxygen voltage sensitive) domains and a kinase domain. Each LOV domain non-covalently binds a flavin mononucleotide (FMN). When we began working on these systems in 2001, the reaction mechanism of all these photoreceptors was unknown.
In the meantime, we have characterized in detail [7-18] the photocycle of two LOV domains, both from the phototropin of the green alga chlamydomonas reinhardtii (C.r.). The chemical nature of two short lived intermediates and the long lived signalling state were identified by transient absorption spectroscopy, employing several point mutants. Since flavoproteins are known to be involved in many redox reactions, a change of the redox state was expected as the signalling event. Quite unexpectedly, however, a reaction through the FMN triplet state was observed, leading to the covalent adduct of FMN to a cystein residue at C4a of the flavin. The details of the mechanism are still under debate.


Left: X-Ray structure of LOV1 of C.r. from [6]; Right: Photocycle of LOV1 of C.r. [7].

Formation of the FMN-cysteine adduct makes the beginning of the signaling chain, which involves activation of a kinase and autophosphorylation of the phototropin. The presence of two different LOV domains in the phototropin suggests that both play a (probably different) role in the signal transduction, and that the interaction between these domains is modified by the adduct formation. Biochemical studies indicate that LOV domains like to form dimers, and that the monomer/dimer ratio changes upon irradiation [16]. When the photoactive cysteine is replaced by glycin, a photoreduction leading to the neutral FMNH* radical is observed as the first reaction following formation of the triplet. The electron donor can be EDTA [15] or an aliphatic mercaptane. With CH3-SH a further intermediate is observed that has the same absorption spectrum as the natural photoadduct [17,18]. Based on these findings, a model for the function of the full phototropin protein is proposed.

In addition to the studies on LOV domains we also performed studies on some cryptochromes [19,20], BLUF proteins, and rhodopsins.

References:

  1. M. A. van der Horst, K. J. Hellingwerf, Acc. Chem. Res. 2004, 37, 13-20.
  2. W. R. Briggs, E. Huala, Annu. Rev. Cell Dev. Biol. 1999, 15, 33-62.
  3. C. Lin, T. Todo, Genome Biology 2005, 6, 220.1-220.9.
  4. M. Gomelsky, G. Klug, Trends Biochem. Sci. 2002, 27, 497-500.
  5. W. R. Briggs, J. M. Christie, Trends Plant Sci. 2002, 7, 204-210.
  6. R. Fedorov, I. Schlichting, E. Hartmann, T. Domratcheva, M. Fuhrmann, P. Hegemann, Biophys. J. 2003, 84, 2474-2482.
  7. T. Kottke, J. Heberle, D. Hehn, B. Dick, P. Hegemann, Phot-LOV1: Photocycle of a blue-light receptor domain from the green alga Chlamydomonas reinhardtii, Biophys. J. 2003, 84, 1192-1201.
  8. T. Kottke, B. Dick, R. Fedorov, I. Schlichting, R. Deutzmann, P. Hegemann, Irreversible photoreduction of flavin in a mutated Phot-LOV1 domain, Biochemistry 2003, 42, 9854-9862.
  9. Huimin Guo, T. Kottke, P. Hegemann, B. Dick, The Phot LOV2 domain and its interaction with LOV1, Biophys. J. 2005, 89 402 - 412.
  10. S.-H. Song, B. Dick, P. Zirak, A. Penzkofer, T. Schiereis, P. Hegemann, Absorption and emission spectroscopic characterisation of combined wildtype LOV1-LOV2 domain of phot from Chlamydomonas reinhardtii, J. Photochem. Photobiol. B 2005, 81, 55-65.
  11. T. Kottke, P. Hegemann, B. Dick, J. Heberle, The photochemistry of the light-, oxygen-, and voltage-sensitive domains in the algal blue light receptor phot, Biopolymers 2006, 82, 373 -378.
  12. S.-H. Song, B. Dick, A. Penzkofer, Photo-induced reduction of flavin mononucleotide in aqueous solutions, Chem. Phys. 2007, 332, 55-65.
  13. S.-H. Song, B. Dick, A. Penzkofer, P. Hegemann, Photo-reduction of flavin mononucleotide to semiquinone form in LOV domain mutants of blue-light receptor phot from Chlamydomonas reinhardtii, J. Photochem. Photobiol. B 2007, 87, 37-48.
  14. A. Penzkofer, A.K. Bansal, S.-H. Song, B. Dick, Fluorescence quenching of flavins by reductive agents, Chem. Phys, 2007, 336, 14-21.
  15. G. Nöll, G. Hauska, P. Hegemann, K. Lanzl, T. Nöll, M. von Sanden-Flohe, B. Dick, Redox properties of LOV domains: Chemical versus photochemical reduction, and influence on the photocycle, ChemBioChem 2007, 8, 2256-2264.
  16. R.J. Kutta, E.S.A. Hofinger, H. Preuss, G. Bernhardt, B. Dick, Blue-light induced interaction of LOV domains from Chlamydomonas reinhardtii, ChemBioChem 2008, 9, 1931-1938.
  17. K. Lanzl, G. Nöll, B. Dick, LOV1 protein from Chlamydomonas reinhardtii is a template for the photoadduct formation of FMN and methylmercaptane, ChemBioChem 2008, 9, 861-864.
  18. Lanzl, K; von Sanden-Flohe, M; Kutta, RJ; Dick, B; Photoreaction of mutated LOV photoreceptor domains from Chlamydomonas reinhardtii with aliphatic mercaptans: implications for the mechanism of wild type LOV. Phys. Chem. Chem. Phys. 12, 6594-6604, 2010.
  19. S.-H. Song, B. Dick, A. Penzkofer, R. Pokorny, A. Batschauer, L.O. Essen, Absorption and fluorescence spectroscopic characterization of cryptochrome 3 from Arabidopsis thaliana, J. Photochem. Photobiol. B 2006, 85, 1-16.
  20. T. Langenbacher, D. Immeln, B. Dick, T. Kottke, Microsecond Light-induced Proton Transfer to Flavin in the Blue Light Sensor Plant Cryptochrome, J. Amer. Chem. Soc. 2009, 131, 14274-14280.
Other publications of B. Dick,


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Address: Institut für Physikalische und Theoretische Chemie, Universitätsstrasse 31, 93053 Regensburg, Germany
Lehrstuhl für Physikalische Chemie - Prof. Dr. Bernhard Dick
Tel: +49 941 943 4486, Fax: +49 941 943 4488, E-Mail: Bernhard.dick[at]chemie.uni-regensburg.de