Photo-CIDNP MAS NMR in Intact Cells ofRhodobactersphaeroidesR26:  Molecular and Atomic Resolution at Nanomolar Concentration

Abstract

Photochemically induced dynamic nuclear polarization (photo-CIDNP) is observed in photosynthetic reaction centers of the carotenoid-less strain R26 of the purple bacterium Rhodobacter sphaeroides by (13)C solid-state NMR at three different magnetic fields (4.7, 9.4, and 17.6 T). The signals of the donor appear enhanced absorptive (positive) and of the acceptor emissive (negative). This spectral feature is in contrast to photo-CIDNP data of reactions centers of Rhodobacter sphaeroides wildtype reported previously (Prakash, S.; Alia; Gast, P.; de Groot, H. J. M.; Jeschke, G.; Matysik, J. J. Am. Chem. Soc. 2005, 127, 14290-14298) in which all signals appear emissive. The difference is due to an additional mechanism occurring in RCs of R26 in the long-living triplet state of the donor, allowing for spectral editing by different enhancement mechanisms. The overall shape of the spectra remains independent of the magnetic field. The strongest enhancement is observed at 4.7 T, enabling the observation of photo-CIDNP enhanced NMR signals from reaction center cofactors in entire bacterial cells allowing for detection of subtle changes in the electronic structure at nanomolar concentration of the donor cofactor. Therefore, we establish in this paper photo-CIDNP MAS NMR as a method to study the electronic structure of photosynthetic cofactors at the molecular and atomic resolution as well as at cellular concentrations.

Photo-CIDNP in the Reaction Center of the Diatom Cyclotella meneghiniana Observed by 13C MAS NMR

“…[61][62] Also entirely emissive spectra have been observed on PS I particles of Spinacia oleracea (spinach). [20] In contrast, RCs of R. sphaeroides R26 [9][62] and PS II (D1/D2 preparation) of spinach [22] show enhanced absorptive signals of the aromatic carbons together with the emissively polarized methine carbons. In these samples, due to the absence of carotenes in the vicinity of the donor, long donor triplet lifetimes, and therefore the DR mechanism can occur.…”

“…[9] The efficiency of the DR mainly depends on the longitudinal relaxation time, which must be in the range of a few hundred nanoseconds or a couple of microseconds. It strongly depends on the Zeeman field B 0 , the temperature and in that temperature range possibly occurring glass transitions.…”

“…[8] Under continuous illumination, up to three spin-chemical mechanisms running in parallel have been revealed contributing to the photo-chemically induced dynamic polarization (photo-CIDNP) in the solid state in photosynthetic RCs: the three spin mixing (TSM), the differential decay (DD) and the differential relaxation (DR) mechanisms. [9][10] [11] [12] Photosynthesis is a photochemical process that is initiated by the light-induced electron transfer in a photosynthetic reaction center (RC). There are two types of photosynthetic RCs that can be found in nature.…”

15N Photo-CIDNP MAS NMR To Reveal Functional Heterogeneity in Electron Donor of Different Plant Organisms

“…In solid-state NMR upon illumination with continuous white light, photo-CIDNP has been observed by applying 15 N magic-angle spinning (MAS) NMR to four photosynthetic RCs: (i) Rhodobacter (R.) sphaeroides R26 [17][18][19][20], wild type (WT) [21][22][23], PSI [2,24], and PSII [24]. Photo-CIDNP is caused by the strong electron polarization in the initial radical pair and subsequently transferred to nuclei, where it is detected by NMR as a signal enhancement up to a factor of about 10,000 [25,26].…”

15 N photochemically induced dynamic nuclear polarization magic-angle spinning NMR analysis of the electron donor of photosystem II

“…The solid-state photo-CIDNP effect, discovered in 1994 (6), relies on different mechanisms called three-spin mixing (7), differential decay (8), and differential relaxation (9, 10). Recently, the contribution of these three mechanisms has been analyzed by field-dependent measurements on unlabeled RCs of the purple bacterium Rhodobacter sphaeroides (11,12). The chemical shift refers to the electronic ground-state after the photocycle, the photo-CIDNP signal intensity is linked to the intermediate radical state.…”