Daniel Thomsson

Organization of Bacteriochlorophylls in Individual Chlorosomes from Chlorobaculum tepidum Studied by 2-Dimensional Polarization Fluorescence Microscopy

Chlorosomes are the largest and most efficient natural light-harvesting systems and contain supramolecular assemblies of bacteriochlorophylls that are organized without proteins. Despite a recent structure determination for chlorosomes from Chlorobaculum tepidum (Ganapathy Proc. Natl. Acad. Sci. U.S.A. 2009, 106, 8525), the issue of a possible large structural disorder is still discussed controversially. We have studied individual chlorosomes prepared under very carefully controlled growth condition by a novel 2-dimensional polarization single molecule imaging technique giving polarization information for both fluorescence excitation and emission simultaneously. Contrary to the existing literature data, the polarization degree or modulation depth (M) for both excitation (absorption) and emission (fluorescence) showed extremely narrow distributions. The fluorescence was always highly polarized with M ≈ 0.77, independent of the excitation wavelength. Moreover, the fluorescence spectra of individual chlorosomes were identical within the error limits. These results lead us to conclude that all chlorosomes possess the same type of internal organization in terms of the arrangement of the bacteriochlorophyll c transition dipole moments and their total excitonic transition dipole possess a cylindrical symmetry in agreement with the previously suggested concentric multitubular chlorophyll aggregate organization (Ganapathy Proc. Natl. Acad. Sci. U.S.A. 2009, 106, 8525).

Oligothiophene Assemblies Defined by DNA Interaction: From Single Chains to Disordered Clusters

The organization of conjugated polyelectrolytes (CPEs) interacting with biomolecules sets conditions for the biodetection of biological processes and identity, through the use of optical emission from the CPE. Herein, a well-defined CPE and its binding to DNA is studied. By using dynamic light scattering and circular dichroism spectroscopy, it is shown that the CPE forms a multimolecule ensemble in aqueous solution that is more than doubled in size when interacting with a small DNA chain, while single chains are evident in ethanol. The related changes in the fluorescence spectra upon polymer aggregation are assigned to oscillator strength redistribution between vibronic transitions in weakly coupled H-aggregates. An enhanced single-molecule spectroscopy technique that allows full control of excitation and emission light polarization is applied to combed and decorated lambdaDNA chains. It is found that the organization of combed CPE-lambdaDNA complexes (when dry on the surface) allows considerable variation of CPE distances and direction relative to the DNA chain. By analysis of the polarization data energy transfer between the polymer chains in individual complexes is confirmed and their sizes estimated.

Correlation Analysis of Fluorescence Intensity and Fluorescence Anisotropy Fluctuations in Single-Molecule Spectroscopy of Conjugated Polymers.

Single-molecule spectroscopy techniques are used to investigate time fluctuations of the fluorescence properties of two different types of conjugated polymer, a polythiophene derivative (PDOPT) and a phenylene vinylene derivative (MEH-PPV), at 100 and 293 K. Linear correlation coefficients between fluorescence intensity and polarization are used to characterize fluctuations. We are able to distinguish between different blinking and bleaching effects on the polarization. Furthermore, the polarization data reveal clear differences in the topology of these two polymers, which is related to the ordered conformation of the MEH-PPV. Plots of correlation coefficients appear to be very different for the two polymers and are also very sensitive to temperature. These observations prove that correlation analysis is a useful tool to understand fluorescence fluctuations in multi-chromophoric systems.

Polarization single complex imaging of circular photosynthetic antenna

Single complex fluorescence polarization spectroscopy is applied to study the peripheral light harvesting antenna (LH2) from photosynthetic purple bacterium Rhodopseudomonas (Rps.) acidophila. The measured two-dimensional excitation-emission polarization plots are used to construct geometric representation for the absorbing B800 and emitting B850 as ellipses. The shape and orientation of the ellipses is discussed in terms of tilted LH2 complexes where emission occurs from energetically disordered B850 excitons.

Excitation polarization provides structural resolution of individual non-blinking nano-objects

We propose to combine the method of fluorescence intensity centroid localization with rotation of the plane of excitation polarization. Polarized light interacts selectively with differently oriented fluorophores; thus yielding topological information on the nanometer scale, without any need for fluorophore blinking. The method is applicable to photostable individual systems, when most of the traditional super-resolution methods fail. A theoretical study is supported by experiments on 30 nm long cyclodextrin-encapsulated single polyrotaxane conjugated polymer chains.