Light-Induced Triplet–Triplet Electron Resonance Spectroscopy

Abstract

We present a new technique, light-induced triplet–triplet electron resonance spectroscopy (LITTER), which measures the dipolar interaction between two photoexcited triplet states, enabling both the distance and angular distributions between the two triplet moieties to be determined on a nanometer scale. This is demonstrated for a model bis-porphyrin peptide that renders dipolar traces with strong orientation selection effects. Using simulations and density functional theory calculations, we extract distance distributions and relative orientations of the porphyrin moieties, allowing the dominant conformation of the peptide in a frozen solution to be identified. LITTER removes the requirement of current light-induced electron spin resonance pulse dipolar spectroscopy techniques to have a permanent paramagnetic moiety, becoming more suitable for in-cell applications and facilitating access to distance determination in unmodified macromolecular systems containing photoexcitable moieties. LITTER also has the potential to enable direct comparison with Förster resonance energy transfer and combination with microscopy inside cells.

Pentacene in 1,3,5-Tri(1-naphtyl)benzene: A Novel Standard for Transient EPR Spectroscopy at Room Temperature

“…Transient or time-resolved EPR (TREPR) [4,14,41] is, therefore, regularly employed in a growing number of laboratories to detect short-lived states including triplets, quintets [35] and radical-pairs [6] in, for example, proteins [43], materials for organic electronics [5,42] or quantum information science [27]. In the field of dipolar spectroscopy and double electron-electron resonance (DEER) photoexcited triplet states offer the possibility of measuring distances in proteins rather than using conventional nitroxide spin-labels (e.g., (2,2,6,6-Tetramethylpiperidin-1-yl)oxyl, TEMPO) [2,3]. From these techniques, further highly specific experimental methods can be derived.…”

Fullerene-based triplet spin labels: methodology aspects for pulsed dipolar EPR spectroscopy

“…16,17 The first work demonstrating the capabilities of dipolar EPR with photoexcited triplet labels was carried out using porphyrins, 16 and several other studies have followed in a few years. [18][19][20][21][22][23][24][25][26][27] The main advantage of porphyrins is their good water solubility, which allows spin labeling 16,20 or complexation with proteins. 23 The first application of triplet porphyrins was demonstrated for the 4-pulse DEER experiment, where porphyrins were photoexcited shortly before the microwave pulse sequence and used as an observer (laser-induced (Li) DEER).…”

Orientation-Selective and Frequency-Correlated Light-Induced Pulsed Dipolar Spectroscopy

“…Recently, we introduced a new triplet-based PDS technique, light-induced triplet–triplet electron resonance spectroscopy (LITTER), which measures the dipolar interaction between two photoexcited triplet states . The benefits of the LITTER experiment were evidenced, and significant orientation selection was demonstrated for a model diporphyrin–peptide system.…”

“…Orientation-dependent simulations were performed using the algorithm previously published, and the results were fitted to the experimental results using an iterative least-squares fitting process, similar to that reported here. , As it is not possible to resolve the orientation of the ZFS tensor from these experiments, in the ReLaserIMD simulations the TPP moiety was modeled as a single point of spin-density located at the center of the porphyrin moiety. In the fitting process the modulation depth was normalized to 1 to account for variations in the measured modulation depths, which may arise from small oscillations in laser power during the experimental acquisition.…”

“…35 Recently, we introduced a new triplet-based PDS technique, light-induced triplet−triplet electron resonance spectroscopy (LITTER), which measures the dipolar interaction between two photoexcited triplet states. 53 The benefits of the LITTER experiment were evidenced, and significant orientation selection was demonstrated for a model diporphyrin−peptide system. In the LITTER experiment, as with all one-dimensional (1D) PDS techniques, multiple traces must be recorded at different fields in order to sample the orientation selection fully.…”