D‐Band EPR and ENDOR Spectroscopy of 15N‐Labeled Photosystem I

Abstract

For billions of years, nature has optimized the photosynthetic machinery that converts light energy into chemical energy. Key primary reactions of photosynthesis occur in large membrane protein-cofactor complexes. The light-induced sequential electron transfer reactions occur through a chain of donor/acceptor cofactors embedded in the protein matrix resulting in a long-lived transmembrane charge-separated state. EPR is the method of choice to study electron transfer and the interaction of protein environment with redox-active cofactors. However, the spectra of organic cofactor radicals typically are not fully resolved and severely overlap at conventional X-band EPR. Even at Q-band EPR, this overlap is present and often a serious problem. As a result, there is a large variation of the reported EPR data and limited understanding of electronic structures of several redox-active cofactors. These serious problems can often be overcome by the excellent spectral resolution provided by high-frequency EPR (HF EPR). Here, we study the electronic structure of the primary electron donor P 700 and the secondary electron acceptor A 1 of Photosystem I (PSI) using 130 GHz (D-band) EPR and Electron-Nuclear-Double-Resonance (ENDOR) spectroscopy. PSI was isotopically labeled with 15 N (I = ½) to avoid quadrupolar interactions in the most abundant nitrogen isotope 14 N (I = 1) and simplify the ENDOR spectra. ENDOR spectroscopy is central for determining the hyperfine coupling of nitrogen atoms of the two chlorophyll molecules comprising oxidized P 700 and the involvement of protein nitrogen atoms with reduced A 1 . While HF ENDOR of A 1 − allows identification of two nitrogen atoms, HF ENDOR of P 700 + still does not permit unique assignment of the recorded hyperfine couplings.

Scrutinizing the Impact of Alternating Electromagnetic Fields on Molecular Features of the Model Plant Arabidopsis thaliana

“…The analytical MF applied in such studies differs in frequency and exceeds the strength of EMF treatment in our study by orders of magnitude. This is valid for all paramagnetic resonance (EPR) and Electron-Nuclear-Double-Resonance (ENDOR) spectroscopy studies [47] where GHz frequency is applied. Nevertheless, these studies reveal the presence of EMF-sensitive structures in photosynthetic organisms.…”