Fluctuating Two-state Light Harvesting In A Photosynthetic Membrane

The mechanism by which light is converted into chemical energy in a natural photosynthetic system has drawn considerable research interest. Using fluorescence spectroscopy and microscopic imaging, we have observed fluctuating intermolecular protein fluorescence resonant energy transfers (FRET) among light-harvesting proteins I and II (LH1 and LH2) in bacterial photosynthetic membranes. Using two-channel, FRET, photon-counting detection and a novel, two-dimensional cross-correlation function amplitude-mapping analysis, we revealed fluorescence intensity and spectral fluctuations of donor (LH2) and acceptor (LH1) fluorescence involving FRET. Our results suggest that there are dynamic coupled and noncoupled states of the light-harvesting protein assemblies in photosynthetic membranes. The light-harvesting complex assembly under ambient conditions and under water involves dynamic intermolecular structural fluctuations that subsequently disturb the degree of energy transfer coupling between proteins in the membrane. Such intrinsic and dynamic heterogeneity of the native photosynthetic membranes, often submerged under the overall thermally induced spectral fluctuations and not observable in an ensemble-averaged measurement, likely plays a critical role in regulating the light-harvesting efficiency of the photosynthetic membranes

Nanosecond time-resolved resonance Raman and ab initio studies of triplet states and radical cations of halobiphenyls and the radicalcations of phenothiazine, promazine, and chloropromazine

published_or_final_versionChemistryDoctoralDoctor of Philosoph

Time-Resolved Resonance Raman Structural Studies of the pB′ Intermediate in the Photocycle of Photoactive Yellow Protein

Time-resolved resonance Raman spectroscopy is used to obtain chromophore vibrational spectra of the pR, pB′, and pB intermediates during the photocycle of photoactive yellow protein. In the pR spectrum, the C(8)–C(9) stretching mode at 998 cm(−1) is ∼60 cm(−1) lower than in the dark state, and the combination of C–O stretching and C(7)H=C(8)H bending at 1283 cm(−1) is insensitive to D(2)O substitution. These results indicate that pR has a deprotonated, cis chromophore structure and that the hydrogen bonding to the chromophore phenolate oxygen is preserved and strengthened in the early photoproduct. However, the intense C(7)H=C(8)H hydrogen out-of-plane (HOOP) mode at 979 cm(−1) suggests that the chromophore in pR is distorted at the vinyl and adjacent C(8)–C(9) bonds. The formation of pB′ involves chromophore protonation based on the protonation state marker at 1174 cm(−1) and on the sensitivity of the COH bending at 1148 cm(−1) as well as the combined C–OH stretching and C(7)H=C(8)H bending mode at 1252 cm(−1) to D(2)O substitution. The hydrogen out-of-plane Raman intensity at 985 cm(−1) significantly decreases in pB′, suggesting that the pR-to-pB′ transition is the stage where the stored photon energy is transferred from the distorted chromophore to the protein, producing a more relaxed pB′ chromophore structure. The C=O stretching mode downshifts from 1660 to 1651 cm(−1) in the pB′-to-pB transition, indicating the reformation of a hydrogen bond to the carbonyl oxygen. Based on reported x-ray data, this suggests that the chromophore ring flips during the transition from pB′ to pB. These results confirm the existence and importance of the pB′ intermediate in photoactive yellow protein receptor activation