15N photo-CIDNP MAS NMR analysis of a bacterial photosynthetic reaction center of Rhodobacter sphaeroides wildtype.

Abstract

The solid-state photochemically induced dynamic nuclear polarization (photo-CIDNP) effect has been studied in a quinone-depleted uniformly (u-)13C,15N-labeled photosynthetic reaction center (RC) protein from purple bacterium Rhodobacter (R.) sphaeroides wild type (WT). As a method for investigation, solid-state 15N NMR under magic-angle spinning (MAS) is applied under both continuous illumination (steady state) and nanosecond-laser flashes (time-resolved). While all previous 15N photo-CIDNP MAS NMR studies on the purple bacterial RC used the carotenoid-less mutant R26, this is the first using WT samples. The absence of further photo-CIDNP mechanisms (compared to R26) and various couplings (compared to 13C NMR experiments on 13C-labeled samples) allows the simplification of the spin-system. We report 15N signals of the three cofactors forming the spin-correlated radical pair (SCRP) and, based on density-functional theory calculations, their assignment. The simulation of photo-CIDNP intensities and time-resolved 15N photo-CIDNP MAS NMR data matches well to the frame of the mechanistic interpretation. Three spin-chemical processes, namely, radical pair mechanism, three spin mixing, and differential decay, generate emissive (negative) 15N polarization in the singlet decay channel and absorptive (positive) polarization in the triplet decay channel of the SCRP. The absorptive 15N polarization of the triplet decay channel is transiently obscured during the lifetime of the triplet state of the carotenoid (3Car); therefore, the observed 15N signals are strongly emissive. Upon decay of 3Car, the transiently obscured polarization becomes visible by reducing the excess of emissive polarization. After the decline of 3Car, the remaining nuclear hyperpolarization decays with nuclear T1 relaxation kinetics.