Elucidation of the mechanism of blue-light reception by vibrational spectroscopy
Zusammenfassung der Projektergebnisse
The mechanism of three different families of flavin-containing blue light receptors were investigated by vibrational spectroscopy. The photoreactions of light-, oxygen-, or voltage-sensitive (LOV) domains from algal and higher plant phototropin and bacterial YtvA were characterized in detail, revealing differences in structure of the reactive cysteine forming a covalent adduct. Signal propagation was studied in full-length YtvA and full-length phototropin, demonstrating that the output domains respond by changes in secondary structure to illumination of the LOV domains. The impact of the chromophore composition of the blue light using flavin (BLUF) domain was analyzed to resolve the impact of the different moieties of the chromophore with respect to their functional significance. Step-scan Fourier transform infrared (FT-IR) spectroscopy with microsecond time resolution was conducted on LOV and BLUF domains, resolving central aspects of signalling state formation and of the concomitant secondary structural changes. Responses of plant cryptochrome to blue light were studied showing the step-wise formation of the signalling state, a flavin neutral radical, by electron transfer and proton transfer. The proton donor was resolved and assigned by infrared spectroscopy. A study on insect cryptochrome highlighted the differences in mechanism to the plant cryptochromes. Taken together, the overview obtained about the different flavin-based sensors demonstrated a much greater variation in the fundamental steps of signalling induced by blue-light illumination of protein-bound flavin as compared to other biological light sensors.
Projektbezogene Publikationen (Auswahl)
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(2004) Chromophore composition of a heterologously expressed BLUF-domain, Photochem. Photobiol. Sci. 3, 1011-1016
W. Laan, T. Bednarz, J. Heberle, and K. J. Hellingwerf
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(2004) Functional variations among LOV domains as revealed by FT-IR difference spectroscopy, Photochem. Photobiol. Sci. 3, 575-579
T. Bednarz, A. Losi, W. Gärtner, P. Hegemann, and J. Heberle
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(2005) Functional characterization of sensory rhodopsin II from Halobacterium salinarum expressed in Escherichia coli. FEBS Lett. 579, 3147-3151
O. S. Mironova, R. G. Efremov, B. Person, J. Heberle, I. L. Budyak, G. Büldt, and R. Schlesinger
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(2005) Vibrational spectroscopy explores the photoreaction of LOV domains, in: Light Sensing in Plants (edited by Wada, M., Simazaki, K., & Iino, M.), Springer, Berlin, pp. 155-161
J. Heberle
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(2006) Blue-light-induced changes in Arabidopsis cryptochrome 1 probed by FTIR difference spectroscopy, Biochemistry 45, 2472-2479
T. Kottke, A. Batschauer, M. Ahmad, and J. Heberle
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(2006) The photochemistry of the light-, oxygen-, and voltage-sensitive domains in the algal blue light receptor phot, Biopolymers 82, 373-378
T. Kottke, P. Hegemann, B. Dick, and J. Heberle
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(2007) A novel photoreaction mechanism for the circadian blue-light photoreceptor Drosophila cryptochrome, J. Biol. Chem. 282, 13011-13021
A. Berndt, T. Kottke, H. Breitkreuz, R. Dvorsky, S. Hennig, M. Alexander, and E. Wolf
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(2007) Blue light induces radical formation and autophosphorylation in the light-sensitive domain of Chlamydomonas cryptochrome. J. Biol. Chem. 282, 21720–21728
D. Immeln, R. Schlesinger, J. Heberle, and T. Kottke
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(2007) Time-resolved FT-IR spectroscopy traces signal relay within the blue-light receptor AppA. ChemPhysChem. 8, 1787-1789
T. Majerus, T. Kottke, W. Laan, K. Hellingwerf, and J. Heberle
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(2009) Microsecond light-induced proton transfer to flavin in the blue light sensor plant cryptochrome, J. Am. Chem. Soc. 131, 14274-14280
T. Langenbacher, D. Immeln, B. Dick, and T. Kottke
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(2009) Time-resolved Fourier transform infrared study on photoadduct formation and secondary structural changes within the phototropin LOV domain. Biophys. J. 96, 1462-1470
A. Pfeifer, T. Majerus, K. Zikihara, D. Matsuoka, S. Tokutomi, J. Heberle, and T. Kottke
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(2010) Blue light induces global and localized conformational changes in the kinase domain of full-length phototropin, Biochemistry 49, 1024-1032
A. Pfeifer, T. Mathes, Y. Lu, P. Hegemann, and T. Kottke
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(2010) Solid-state photo- CIDNP effect observed in phototropin LOV1-C57S by 13C magic-angle spinning NMR spectroscopy, J. Am. Chem. Soc.
S. S. Thamarath, J. Heberle, P. Hore, T. Kottke, and J. Matysik
