Phytochromes are a diverse family of bilin-binding photoreceptors that regulate a wide range of physiological processes. Their photochemical properties make them attractive for applications in optogenetics and superresolution microscopy. Phytochromes undergo reversible photoconversion triggered by theZ⇄Ephotoisomerization about the double bond in the bilin chromophore. However, it is not fully understood at the molecular level how the protein framework facilitates the complex photoisomerization dynamics. We have studied a single-domain bilin-binding photoreceptor All2699g1 (Nostocsp. PCC 7120) that exhibits photoconversion between the red light-absorbing (Pr) and far red-absorbing (Pfr) states just like canonical phytochromes. We present the crystal structure and examine the photoisomerization mechanism of the Prform as well as the formation of the primary photoproduct Lumi-R using time-resolved spectroscopy and hybrid quantum mechanics/molecular mechanics simulations. We show that the unusually long excited state lifetime (broad lifetime distribution centered at ∼300 picoseconds) is due to the interactions between the isomerizing pyrrole ring D and an adjacent conserved Tyr142. The decay kinetics shows a strongly distributed character which is imposed by the nonexponential protein dynamics. Our findings offer a mechanistic insight into how the quantum efficiency of the bilin photoisomerization is tuned by the protein environment, thereby providing a structural framework for engineering bilin-based optical agents for imaging and optogenetics applications.

Under environmental duress, many organisms accumulate large amounts of osmolytes - molecularly small organic solutes. Osmolytes are known to counteract stress, driving proteins to their compact native states by their exclusion from protein surfaces. In contrast, the effect of osmolytes on lipid membranes is poorly understood and widely debated. Many fully membrane-permeable osmolytes exert an apparent attractive force between lipid membranes, yet all proposed models fail to fully account for the origin of this force. We follow the quintessential osmolyte trimethylamine N-oxide (TMAO) and its interaction with dimyristoyl phosphatidylcholine (DMPC) membranes in aqueous solution. We find that by partitioning away from the inter-bilayer space, TMAO pushes adjacent membranes closer together. Experiments and simulations further show that the partitioning of TMAO away from the volume between bilayers stems from its exclusion from the lipid-water interface, similar to the mechanism of protein stabilization by osmolytes. We extend our analysis to show that the preferential interaction of other physiologically relevant solutes (including sugars and DMSO) also correlates with their effect on membrane bilayer interactions. Our study resolves a long-standing puzzle, explaining how osmolytes can increase membrane-membrane attraction or repulsion depending on their preferential interactions with lipids.

This perspective article highlights the challenges in the theoretical description of photoreceptor proteins using multiscale modeling, as discussed at the CECAM workshop in Tel Aviv, Israel. The participants have identified grand challenges and discussed the development of new tools to address them. Recent progress in understanding representative proteins such as green fluorescent protein, photoactive yellow protein, phytochrome, and rhodopsin is presented, along with methodological developments.