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International audience; The activation of rhodopsin, the light-sensitive G-protein-coupled receptor responsible for dim-light vision in vertebrates, is driven by an ultrafast excited-state double-bond isomerization with a quantum efficiency of almost 70%. The origin of such light sensitivity is not understood and a key question is whether in-phase nuclear motion controls the quantum efficiency value. In this study we used hundreds of quantum-classical trajectories to show that, 15 fs after light absorption, a degeneracy between the reactive excited state and a neighbouring state causes the splitting of the rhodopsin population into subpopulations. These subpopulations propagate with different velocities and lead to distinct contributions to the quantum efficiency. We also show here that such splitting is modulated by protein electrostatics, thus linking amino acid sequence variations to quantum efficiency modulation. Finally, we discuss how such a linkage that in principle could be exploited to achieve higher quantum efficiencies would simultaneously increase the receptor thermal noise leading to a trade-off that may have played a role in rhodopsin evolution.

The use of light-responsive proteins to control both living or synthetic cells, is at the core of the expanding fields of optogenetics and synthetic biology. It is thus apparent that a richer reaction toolbox for the preparation of such systems is of fundamental importance. Here, we provide a proof-of-principle demonstration that Morita-Baylis-Hillman adducts can be employed to perform a facile site-specific, irreversible and diastereoselective click-functionalization of a lysine residue buried into a lipophilic binding pocket and yielding an unnatural chromophore with an extended π-system. In doing so we effectively open the path to the in vitro preparation of a library of synthetic proteins structurally reminiscent of xanthopsin eubacterial photoreceptors. We argue that such a library, made of variable unnatural chromophores inserted in an easy-to-mutate and crystallize retinoic acid transporter, significantly expand the scope of the recently introduced rhodopsin mimics as both optogenetic and "lab-on-a-molecule" tools.

Recently, progress in IR sources has led to the discovery that humans can detect infrared (IR) light. This is hypothesized to be due to the two-photon absorption (TPA) events promoting the retina dim-light rod photoreceptor rhodopsin to the same excited state populated via one-photon absorption (OPA). Here, we combine quantum mechanics/molecular mechanics and extended multiconfiguration quasi-degenerate perturbation theory calculations to simulate the TPA spectrum of bovine rhodopsin (Rh) as a model for the human photoreceptor. The results show that the TPA spectrum of Rh has an intense S0 → S1 band but shows also S0 → S2 and S0 → S3 transitions whose intensities, relative to the S0 → S1 band, are significantly increased when compared to the corresponding bands of the OPA spectrum. In conclusion, we show that IR light in the 950 nm region can be perceived by rod photoreceptors, thus supporting the two-photon origin of the IR perception. We also found that the same photoreceptor can perceive red (i.e., close to 680 nm) light provided that TPA induces population of S2.

This is the peer reviewed version of the following article: Mroginski, Adam, Amoyal, Barnoy, Bondar, ... Schapiro. (2020). Frontiers in Multiscale Modeling of Photoreceptor Proteins. Photochemistry and Photobiology, 97(2):243-269, which has been published in final form at https://doi.org/10.1111/php.13372. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. This article may not be enhanced, enriched or otherwise transformed into a derivative work, without express permission from Wiley or by statutory rights under applicable legislation. Copyright notices must not be removed, obscured or modified. The article must be linked to Wiley’s version of record on Wiley Online Library and any embedding, framing or otherwise making available the article or pages thereof by third parties from platforms, services and websites other than Wiley Online Library must be prohibited. 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.

The automatic building of quantum mechanical/molecular mechanical models (QM/MM) of rhodopsins has been recently proposed. This is a prototype of an approach that will be expanded to make possible the systematic computational investigation of biological photoreceptors. QM/MM models represent useful tools for biophysical studies and for protein engineering, but have the disadvantage of being time-consuming to construct, error prone and, also, of not being consistently constructed by researchers operating in different laboratories. These basic issues impair the possibility to comparatively study hundreds of photoreceptors of the same family, as typically required in biological or biotechnological studies. Thus, in order to carry out systematic studies of photoreceptors or, more generally, light-responsive proteins, some of the authors have recently developed the Automatic Rhodopsin Modeling (ARM) protocol for the fast generation of combined QM/MM models of photoreceptors formed by a single protein incorporating in its cavity a single chromophore. In this chapter, we review the results of such research effort by revising the building protocol and benchmark studies and by discussing selected applications.

This article introduces Web-ARM, a specialized tool, online available, designed to build quantum mechanical/molecular mechanical models of rhodopsins, a widely spread family of light-responsive proteins. Web-ARM allows the rapidly building of models of rhodopsins with a documented quality and the prediction of trends in UV-vis absorption maximum wavelengths, based on their excitation energies computed at the CASPT2//CASSCF/Amber level of theory. Web-ARM builds upon the recently reported, python-based a-ARM protocol [J. Chem. Theory Comput., 2019, 15, 3134-3152] and, as such, necessitates only a crystallographic structure or a comparative model in PDB format and a very basic knowledge of the studied rhodopsin system. The user-friendly web interface uses such input to generate congruous, gas-phase models of rhodopsins and, if requested, their mutants. We present two possible applications of Web-ARM, which showcase how the interface can be employed to assist both research and educational activities in fields at the interface between chemistry and biology. The first application shows how, through Web-ARM, research projects (e.g., rhodopsin and rhodopsin mutant screening) can be carried out in significantly less time with respect to using the required computational photochemistry tools via a command line. The second application documents the use of Web-ARM in a real-life educational/training activity, through a hands-on experience illustrating the concepts of rhodopsin color tuning.

Rhodopsins hosting synthetic retinal protonated Schiff base analogues are important for developing tools for optogenetics and high-resolution imaging. The ideal spectroscopic properties of such analogues include long-wavelength absorption/emission and fast/hindered photoisomerization. While the former may be achieved, for instance, by elongating the chromophore π-system, the latter requires a detailed understanding of the substituent effects (i.e. steric or electronic) on the chromophore light-induced dynamics. Below, we compare the results of QM/MM excited state trajectories of native and analogue-hosting microbial rhodopsins from the eubacterium Anabaena. The results uncover a relationship between nature of the substituent on the analogue (i.e. electron donating (a Me group) or withdrawing (a CF3 group)) and rhodopsin excited state lifetime. Most importantly, we show that electron donating or With drawing substituents cause a decrease or an increase in the electronic mixing of the first two excited states which, in turn, controls the photoisomerization speed.

While the light-induced population dynamics of different photoresponsive proteins has been investigated spectroscopically, systematic computational studies have not yet been possible due to the phenomenally high cost of suitable high level quantum chemical methods and the need of propagating hundreds, if not thousands, of nonadiabatic trajectories. Here we explore the possibility of studying the photodynamics of rhodopsins by constructing and investigating quantum mechanics/molecular mechanics (QM/MM) models featuring reduced retinal chromophores. In order to do so we use the sensory rhodopsin found in the cyanobacterium Anabaena PCC7120 (ASR) as a benchmark system. We find that the basic mechanistic features associated with the excited state dynamics of ASR QM/MM models are reproduced using models incorporating a minimal (i.e., three double-bond) chromophore. Furthermore, we show that ensembles of nonadiabatic ASR trajectories computed using the same abridged models replicate, at both the CASPT2 and CASSCF levels of theory, the trends in spectroscopy and lifetimes estimated using unabridged models and observed experimentally at room temperature. We conclude that a further expansion of these studies may lead to low-cost QM/MM rhodopsin models that may be used as effective tools in high-throughput in silico mutant screening.

Vibronic coupling is key to efficient energy flow in molecular systems and a critical component of most mechanisms invoking quantum effects in biological processes. Despite increasing evidence for coherent coupling of electronic states being mediated by vibrational motion, it is not clear how and to what degree properties associated with vibrational coherence such as phase and coupling of atomic motion can impact the efficiency of light-induced processes under natural, incoherent illumination. Here, we show that deuteration of the H11–C11=C12–H12 double-bond of the 11-cis retinal chromophore in the visual pigment rhodopsin significantly and unexpectedly alters the photoisomerization yield while inducing smaller changes in the ultrafast isomerization dynamics assignable to known isotope effects. Combination of these results with non-adiabatic molecular dynamics simulations reveals a vibrational phase-dependent isotope effect that we suggest is an intrinsic attribute of vibronically coherent photochemical processes.

Spectral data show that the photoisomerization of retinal protonated Schiff base (rPSB) chromophores occurs on a 100 fs time scale or less in vertebrate rhodopsins, it is several times slower in microbial rhodopsins and it is between one and 2 orders of magnitude slower in solution. These time scale variations have been attributed to specific modifications of the topography of the first excited state potential energy surface of the chromophore. However, it is presently not clear which specific environment effects (e.g., electrostatic, electronic, or steric) are responsible for changing the surface topography. Here, we use QM/MM models and excited state trajectory computations to provide evidence for an increase in electronic mixing between the first and the second excited state of the chromophore when going from vertebrate rhodopsin to the solution environments. Ultimately, we argue that a correlation between the lifetime of the first excited state and electronic mixing between such state and its higher neighbor, may have been exploited to evolve rhodopsins toward faster isomerization and, possibly, light-sensitivity.

We report on a prototype protocol for the automatic and fast construction of congruous sets of QM/MM models of rhodopsin-like photoreceptors and of their mutants. In the present implementation the information required for the construction of each model is essentially a crystallographic structure or a comparative model complemented with information on the protonation state of ionizable side chains and distributions of external counterions. Starting with such information, a model formed by a fixed environment system, a flexible cavity system, and a chromophore system is automatically generated. The results of the predicted vertical excitation energy for 27 different rhodopsins including vertebrate, invertebrate, and microbial pigments indicate that such basic models could be employed for predicting trends in spectral changes and/or correlate the spectral changes with structural variations in large sets of proteins.