Photosynthetic reaction center variants made via genetic code expansion show Tyr at M210 tunes the initial electron transfer mechanism

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Abstract

Photosynthetic reaction centers (RCs) from Rhodobacter sphaeroides were engineered to vary the electronic properties of a key tyrosine (M210) close to an essential electron transfer component via its replacement with site-specific, genetically encoded noncanonical amino acid tyrosine analogs. High fidelity of noncanonical amino acid incorporation was verified with mass spectrometry and X-ray crystallography and demonstrated that RC variants exhibit no significant structural alterations relative to wild type (WT). Ultrafast transient absorption spectroscopy indicates the excited primary electron donor, P*, decays via a ∼4-ps and a ∼20-ps population to produce the charge-separated state P ⁺H _A ⁻ in all variants. Global analysis indicates that in the ∼4-ps population, P ⁺H _A ⁻ forms through a two-step process, P*→ P ⁺B _A ⁻→ P ⁺H _A ⁻, while in the ∼20-ps population, it forms via a one-step P* → P ⁺H _A ⁻ superexchange mechanism. The percentage of the P* population that decays via the superexchange route varies from ∼25 to ∼45% among variants, while in WT, this percentage is ∼15%. Increases in the P* population that decays via superexchange correlate with increases in the free energy of the P ⁺B _A ⁻ intermediate caused by a given M210 tyrosine analog. This was experimentally estimated through resonance Stark spectroscopy, redox titrations, and near-infrared absorption measurements. As the most energetically perturbative variant, 3-nitrotyrosine at M210 creates an ∼110-meV increase in the free energy of P ⁺B _A ⁻ along with a dramatic diminution of the 1,030-nm transient absorption band indicative of P ⁺B _A ^– formation. Collectively, this work indicates the tyrosine at M210 tunes the mechanism of primary electron transfer in the RC.

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Global and target analysis of time-resolved spectra.

Ivo H M van Stokkum, Delmar S Larsen, Rienk van Grondelle (2004)

In biological/bioenergetics research the response of a complex system to an externally applied perturbation is often studied. Spectroscopic measurements at multiple wavelengths are used to monitor the kinetics. These time-resolved spectra are considered as an example of multiway data. In this paper, the methodology for global and target analysis of time-resolved spectra is reviewed. To fully extract the information from the overwhelming amount of data, a model-based analysis is mandatory. This analysis is based upon assumptions regarding the measurement process and upon a physicochemical model for the complex system. This model is composed of building blocks representing scientific knowledge and assumptions. Building blocks are the instrument response function (IRF), the components of the system connected in a kinetic scheme, and anisotropy properties of the components. The combination of a model for the kinetics and for the spectra of the components results in a more powerful spectrotemporal model. The model parameters, like rate constants and spectra, can be estimated from the data, thus providing a concise description of the complex system dynamics. This spectrotemporal modeling approach is illustrated with an elaborate case study of the ultrafast dynamics of the photoactive yellow protein. Copyright 2004 Elsevier B.V.

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Synthetic Methods Driven by the Photoactivity of Electron Donor–Acceptor Complexes

Giacomo E. M. Crisenza, Daniele Mazzarella, Paolo Melchiorre (2020)

The association of an electron-rich substrate with an electron-accepting molecule can generate a new molecular aggregate in the ground state, called an electron donor–acceptor (EDA) complex. Even when the two precursors do not absorb visible light, the resulting EDA complex often does. In 1952, Mulliken proposed a quantum-mechanical theory to rationalize the formation of such colored EDA complexes. However, and besides a few pioneering studies in the 20th century, it is only in the past few years that the EDA complex photochemistry has been recognized as a powerful strategy for expanding the potential of visible-light-driven radical synthetic chemistry. Here, we explain why this photochemical synthetic approach was overlooked for so long. We critically discuss the historical context, scientific reasons, serendipitous observations, and landmark discoveries that were essential for progress in the field. We also outline future directions and identify the key advances that are needed to fully exploit the potential of the EDA complex photochemistry.