Blue light induces radical formation and autophosphorylation in the light-sensitive domain of Chlamydomonas cryptochrome

Immeln D, Schlesinger R, Heberle J, Kottke T. Blue light induces radical formation and autophosphorylation in the light-sensitive domain of Chlamydomonas cryptochrome. JOURNAL OF BIOLOGICAL CHEMISTRY. 2007;282(30):21720-21728

Infrared spectrum and absorption coefficient of the cofactor flavin in water

Spexard M, Immeln D, Thöing C, Kottke T. Infrared spectrum and absorption coefficient of the cofactor flavin in water. VIBRATIONAL SPECTROSCOPY. 2011;57(2):282-287

Microsecond Light-induced Proton Transfer to Flavin in the Blue Light Sensor Plant Cryptochrome

Langenbacher T, Immeln D, Dick B, Kottke T. Microsecond Light-induced Proton Transfer to Flavin in the Blue Light Sensor Plant Cryptochrome. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY. 2009;131(40):14274-14280

Primary events in the blue light sensor plant cryptochrome: intraprotein electron and proton transfer revealed by femtosecond spectroscopy

Immeln D, Weigel A, Kottke T, Pérez Lustres JL. Primary events in the blue light sensor plant cryptochrome: intraprotein electron and proton transfer revealed by femtosecond spectroscopy. Journal of the American Chemical Society. 2012;134(30):12536-12546.Photoreceptors are chromoproteins that undergo fast conversion from dark to signaling states upon light absorption by the chromophore. The signaling state starts signal transduction in vivo and elicits a biological response. Therefore, photoreceptors are ideally suited for the analysis of protein activation by time-resolved spectroscopy. We focus on plant cryptochromes which are blue light sensors regulating the development and daily rhythm of plants. The signaling state of these flavoproteins is the neutral radical of the flavin chromophore. It forms on the microsecond timescale after light absorption by the oxidized state. We apply here femtosecond broadband transient absorption to early stages of signaling-state formation in an algal plant cryptochrome. Transient spectra show: i) sub-ps decay of flavin stimulated emission and ii) further decay of signal until 100 ps delay with nearly constant spectral shape.i) monitors electron transfer from a nearby tryptophan to the flavin and occurs with a time constant of τ(ET)=0.4 ps. ii) is analyzed by spectral decomposition and occurs with a characteristic time constant τ(1)=31 ps. We reason that hole transport through a tryptophan triad to the protein surface and partial deprotonation of tryptophan cation radical hide behind τ(1). These processes are probably governed by vibrational cooling. Spectral decomposition is used together with anisotropy to obtain the relative orientation of flavin and the final electron donor. This narrows the number of possible electron donors down to two tryptophans. Structural analysis suggests that a set of histidines surrounding the terminal tryptophan may act as proton acceptor and thereby stabilize the radical pair on a 100 ps timescale

Photoreaction of Plant and DASH Cryptochromes Probed by Infrared Spectroscopy: The Neutral Radical State of Flavoproteins

Immeln D, Pokorny R, Herman E, Moldt J, Batschauer A, Kottke T. Photoreaction of Plant and DASH Cryptochromes Probed by Infrared Spectroscopy: The Neutral Radical State of Flavoproteins. Journal of Physical Chemistry B. 2010;114(51):17155-17161

Primary Events in the Blue Light Sensor Plant Cryptochrome: Intraprotein Electron and Proton Transfer Revealed by Femtosecond Spectroscopy

Photoreceptors are chromoproteins that undergo fast conversion from dark to signaling states upon light absorption by the chromophore. The signaling state starts signal transduction in vivo and elicits a biological response. Therefore, photoreceptors are ideally suited for analysis of protein activation by time-resolved spectroscopy. We focus on plant cryptochromes which are blue light sensors regulating the development and daily rhythm of plants. The signaling state of these flavoproteins is the neutral radical of the flavin chromophore. It forms on the microsecond time scale after light absorption by the oxidized state. We apply here femtosecond broad-band transient absorption to early stages of signaling-state formation in a plant cryptochrome from the green alga <i>Chlamydomonas reinhardtii</i>. Transient spectra show (i) subpicosecond decay of flavin-stimulated emission and (ii) further decay of signal until 100 ps delay with <i>nearly</i> constant spectral shape. The first decay (i) monitors electron transfer from a nearby tryptophan to the flavin and occurs with a time constant of τ_ET = 0.4 ps. The second decay (ii) is analyzed by spectral decomposition and occurs with a characteristic time constant τ₁ = 31 ps. We reason that hole transport through a tryptophan triad to the protein surface and partial deprotonation of tryptophan cation radical hide behind τ₁. These processes are probably governed by vibrational cooling. Spectral decomposition is used together with anisotropy to obtain the relative orientation of flavin and the final electron donor. This narrows the number of possible electron donors down to two tryptophans. Structural analysis suggests that a set of histidines surrounding the terminal tryptophan may act as proton acceptor and thereby stabilize the radical pair on a 100 ps time scale