Light is vital for all forms of life. Living organisms use biological light sensors, known as photoreceptors, to react to light. But how these molecular light sensors work is not yet understood in full detail. A research team including UniSysCat researchers Andrea Schmidt, Peter Hildebrandt and Patrick Scheerer in cooperation with the UniSysCat SAB member Junko Yano has been able to gain very precise insights into the light-controlled and time-dependent structural changes of a photoreceptor and thus its molecular function.
Organisms use light to regulate many important physiological processes from vision to the “internal clock” and the expression of genes. To perceive light, organisms use special proteins, so-called photoreceptors. These carry a certain light-sensing molecule, the chromophore, which changes its structure when the light conditions change and thus acts like a switch.
These structural changes are set in motion by exposure to light and are orchestrated with great precision. Not only the chromophore is involved here, but also its environment in the photoreceptor plays an important role: the motion of the chromophore is precisely coupled with structural changes in surrounding protein parts. A complex spectacle that nature has developed here.
In their recent study, the research team investigated how this coupling is achieved in a bacterial photoreceptor, a variant of the phytochrome Agp2 from the soil bacterium Agrobacterium fabrum, which is sensitive to dark-red light. Using serial-femtosecond crystallography and pump-probe techniques at two free electron laser sources, Linac Coherent Light Source (LCLS; Stanford, USA) and SPring-8 Angstrom Compact free electron LAser (SACLA; Kouto, Japan), the researchers collected a series of structural snapshots of the moving photoreceptor at different times after illumination and incubation. The chosen method has several advantages over classical protein crystallography: For example, intermediate states of photoreceptor movements can be captured under room temperature conditions, whereas classical protein crystallography usually only visualizes (cryogenically frozen) stable ground states. Based on selected structural data, the initial photoisomerization of the chromophore was also calculated using hybrid quantum mechanics/molecular mechanics (QM/MM) simulations to complement the experimental data
In this way, the research team was able to observe the structural movements of the protein in response to light, just like in a flip-book: after the initial photoisomerization of the chromophore, the chromophore and its immediate surroundings in the photoreceptor first relax. Then the amino acids in the chromophore binding pocket adapt structurally, which in turn induces the conformational switch to initiate the restructuring of a close-lying protein peptide segment called the "tongue". These events allow the identification of the molecular switch that is linked to an intramolecular proton transfer as a prerequisite for the subsequent structural transitions.
Thus, the present series of structural snapshots allows a particularly detailed description of the sequence of structural changes after photoisomerization of a photoreceptor. The research team was able to shed light on the coupling of the motion of the chromophore with the structural changes in its environment - a crucial step in understanding the (de)activation process of this photoreceptor. These results are impressive in themselves, but they also clearly show how powerful the chosen conceptual and methodological approach is for understanding the functioning of photoreceptors in general.
This study has been published in Science Advances: Luisa Sauthof et al., Serial-femtosecond crystallography reveals how a phytochrome variant couples chromophore and protein structural changes. Sci. Adv. 11, eadp2665 (2025). DOI:10.1126/sciadv.adp2665