Ultrafast Charge-Transfer Dynamics in the Iron–Sulfur Complex of <i>Rhodobacter capsulatus</i> Ferredoxin VI

Iron–sulfur proteins play essential roles in various biological processes. Their electronic structure and vibrational dynamics are key to their rich chemistry but nontrivial to unravel. Here, the first ultrafast transient absorption and impulsive coherent vibrational spectroscopic (ICVS) studies on 2Fe–2S clusters in <i>Rhodobacter capsulatus</i> ferreodoxin VI are characterized. Photoexcitation initiated populations on multiple excited electronic states that evolve into each other in a long-lived charge-transfer state. This suggests a potential light-induced electron-transfer pathway as well as the possibility of using iron–sulfur proteins as photosensitizers for light-dependent enzymes. A tyrosine chain near the active site suggests potential hole-transfer pathways and affirms this electron-transfer pathway. The ICVS data revealed vibrational bands at 417 and 484 cm^–1, with the latter attributed to an excited-state mode. The temperature dependence of the ICVS modes suggests that the temperature effect on protein structure or conformational heterogeneities needs to be considered during cryogenic temperature studies

Excited-State Self-Trapping and Ground-State Relaxation Dynamics in Poly(3-hexylthiophene) Resolved with Broadband Pump–Dump–Probe Spectroscopy

Broadband femtosecond transient absorption spectroscopy is used to explore the mechanisms underlying excited-state and ground-state exciton relaxation in poly(3-hexylthiophene) (P3HT) solution. We focus on the picosecond spectral shifts in the ground and excited states of P3HT, using pump–probe (PP) and pump–dump–probe (PDP) techniques to investigate exciton relaxation mechanisms. Excited-state PP signals resolved a dynamic stimulated emission Stokes shift and ground-state reorganization; PDP signals resolved a blue-shifting nonequilibrium ground-state bleach. Initial structural reorganization is shown to be faster in the excited state. Ground-state reorganization is shown to be dependent on dump time, with later times resulting in relatively more population undergoing slow (∼20 ps) reorganization. These observations are discussed in the context of structural relaxation involving small-scale (<1 ps) and large-scale (>1 ps) planarization of thiophene groups following photoexcitation. Excited-state and ground-state dynamics are contrasted in terms of electronic structure defining the torsional potential energy surfaces. It is shown that the primary excitonic relaxation mechanism is excited-state self-trapping via torsional relaxation rather than exciton energy transfer

Subpicosecond Excited-State Proton Transfer Preceding Isomerization During the Photorecovery of Photoactive Yellow Protein

The ultrafast excited-state dynamics underlying the receptor state photorecovery is resolved in the M100A mutant of the photoactive yellow protein (PYP) from Halorhodospira halophila. The M100A PYP mutant, with its distinctly slower photocycle than wt PYP, allows isolation of the pB signaling state for study of the photodynamics of the protonated chromophore <i>cis-p</i>-coumaric acid. Transient absorption signals indicate a subpicosecond excited-state proton-transfer reaction in the pB state that results in chromophore deprotonation prior to the cis−trans isomerization required in the photorecovery dynamics of the pG state. Two terminal photoproducts are observed, a blue-absorbing species presumed to be deprotonated <i>trans-p</i>-coumaric acid and an ultraviolet-absorbing protonated photoproduct. These two photoproducts are hypothesized to originate from an equilibrium of open and closed folded forms of the signaling state, I₂ and I₂′

Excitation-Wavelength-Dependent Photocycle Initiation Dynamics Resolve Heterogeneity in the Photoactive Yellow Protein from <i>Halorhodospira halophila</i>

Photoactive yellow proteins (PYPs) make up a diverse class of blue-light-absorbing bacterial photoreceptors. Electronic excitation of the <i>p</i>-coumaric acid chromophore covalently bound within PYP results in triphasic quenching kinetics; however, the molecular basis of this behavior remains unresolved. Here we explore this question by examining the excitation-wavelength dependence of the photodynamics of the PYP from <i>Halorhodospira halophila</i> via a combined experimental and computational approach. The fluorescence quantum yield, steady-state fluorescence emission maximum, and cryotrapping spectra are demonstrated to depend on excitation wavelength. We also compare the femtosecond photodynamics in PYP at two excitation wavelengths (435 and 475 nm) with a dual-excitation-wavelength-interleaved pump–probe technique. Multicompartment global analysis of these data demonstrates that the excited-state photochemistry of PYP depends subtly, but convincingly, on excitation wavelength with similar kinetics with distinctly different spectral features, including a shifted ground-state beach and altered stimulated emission oscillator strengths and peak positions. Three models involving multiple excited states, vibrationally enhanced barrier crossing, and inhomogeneity are proposed to interpret the observed excitation-wavelength dependence of the data. Conformational heterogeneity was identified as the most probable model, which was supported with molecular mechanics simulations that identified two levels of inhomogeneity involving the orientation of the R52 residue and different hydrogen bonding networks with the <i>p</i>-coumaric acid chromophore. Quantum calculations were used to confirm that these inhomogeneities track to altered spectral properties consistent with the experimental results