Spectroscopic Properties of Photosystems Revealed by Single-molecule Spectroscopy at Low Temperature

Abstract

In this thesis, single-molecule spectroscopy (SMS) at low temperature is utilized to comprehensively investigate the spectroscopic properties of the light-harvesting antenna complexes such as photosystem I (PSI) and II (PSII). The fluorescence of single PSII core complexes (PSIIcc) shows that the spectra are dominated by sharp lines, which come from multiple emitters and not only from one lowest trap. These datasets show the existence of the three well-known fluorescence bands denoted F685, F689, and F695. Further analysis of dimeric PSIIcc (dPSIIcc) presents that these sharp lines are the result of weak to intermediate exciton-vibrational coupling and slow spectral di usion. The Huang-Rhys factors, which are a measure of the strength of exciton-vibrational coupling, vary between 0.03 and 0.8 in single dPSII. Based on these values, there is no obvious correlation between coupling strength and wavelength position. These results show that electrostatic, rather than exchange or dispersive interactions, are the main contributors to the exciton-vibrational coupling in dPSII systems. The SMS of single monomeric PSII core complexes (mPSIIcc) at 1.6 K, also shows the same three fluorescence bands detected in dPSIIcc, however, the intensity of F695 in mPSIIcc SMS datasets is reduced as compared to the single dPSIIcc. The mPSIIcc contains one Beta-Carotene less than dPSIIcc at the monomer-monomer interface of dPSIIcc, which leads to an increased lifetime of the triplet state by Chl 17. This explains the reduced singlet emission of F695 in mPSIIcc. The results from fluorescence measurements of single PSIIcc and PSI complexes show different spectroscopic properties. The fluorescence of single PSI complexes are mostly characterized by sharp lines and a broad emission at 1.6 K. Polarization dependent SMS provides detailed insight into the fluorescence dynamics of the red Chls, demonstrating that Chls absorb light at longer wavelengths than reaction center, and their interactions with other chromophores are mainly happened by excitation energy transfer (EET). The spectral dynamics between the single PSI trimer (PSI-T) and monomer (PSI-M) complexes in both S- and P-polarization dependent datasets indicate that the single PSI-M complexes feature less spectral diversity of their fluorescence than single PSI-Ts. The analysis of polarization dependent datasets from single PSI-M complexes demonstrates two spectrally separate emissions corresponding to two fluorescence pools in PSI-M. Representatively, an almost perpendicular orientation is estimated between them, where they are not connected with each other via energy transfer pathways. Finally, suitable candidates for red-coupled Chl dimers based on the X-ray structure of PSI-M are discussed. The fluorescence enhancements of PSI-T in proximity to bimetallic plasmonic nanostructures are reported using SMS at cryogenic temperature (1.6 K). Moreover, controlled modifcation of fluorescence and energy transfer properties of PSI complexes are shown by using a Fabry-Perot resonator with silver mirrors. Finally, resolution enhancement of a confocal scanning microscope using immersion liquid under cryogenic conditions for imaging the biological specimens is discussed.