PUB - Publikationen an der Universität Bielefeld

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.