Ensemble and single-molecule spectroscopy demonstrates that both emission and absorption of peridinin−chlorophyll−protein photosynthetic antennae can be largely enhanced through plasmonic interactions. We find up to 18-fold increase of the chlorophyll fluorescence for complexes placed near a silver metal layer. This enhancement, which leaves no measurable effects on the protein structure, is observed when exciting either chlorophyll or carotenoid and is attributed predominantly to an increase of the excitation rate in the antenna. The enhancement mechanism comes from plasmon-induced amplification of electromagnetic fields inside the complex. This result is an important step toward applying plasmonic nanostructures for controlling the optical response of complex biomolecules and improving the design and functioning of artificial light-harvesting systems.Strong enhancement of electromagnetic fields generated through plasmon resonances in metal films and particles has recently stimulated a considerable interest in diverse research fields such as optical spectroscopy, cell imaging, quantum information processing, nanophotonics, and biosensors. [1][2][3][4][5] This versatility results from a dramatic influence that plasmons impose on the absorption and emission properties of nearby located dipoles, for example, semiconductor nanocrystals and nanowires 6-12 or dye molecules. [13][14][15][16][17][18] Optical response of an emitter coupled to a plasmonic structure depends upon spatial arrangement as well as spectral characteristics of a studied system. Remarkable progress has been made in on-demand design of metal nanostructures, which is essential for tuning the resonance frequency and thus the coupling strength. 13,14,19 Complementary efforts focused on developing advanced experiments to study dipoles placed in the vicinity of a metal nanoparticle have shed light on the interplay between radiative and nonradiative processes in these systems. 16,18 This very relation determines whether the fluorescence is enhanced [9][10][11]16 or quenched due to the dominating role of nonradiative energy transfer from the dipole to the metal. 15,18 Metal-enhanced fluorescence (MEF) has been observed for many hybrid systems that include nanocrystals on corrugated metal surfaces, 10,11 dye molecules coupled to metal nanoparticles, 18 and nanocrystal-nanoparticle bioconjugates. 8 In all these cases, very stable and highly fluorescing emitters have been selected. It would be, however, highly desirable to apply MEF to weakly fluorescing systems such as DNA, 20 carbon nanotubes 21 or, yet experimentally unexplored in this context, light-harvesting complexes. These latter proteinpigment systems, which contain chlorophyll (Chl) and carotenoid molecules embedded in a protein matrix, participate in the photosynthesis process by collecting sunlight energy and transferring it to reactions centers. The presence of fluorescing Chls and the protein, separated by a few nanometers, renders light-harvesting complexes ideal for studying the prot...

Single molecule spectroscopy experiments are reported for native peridinin-chlorophyll a-protein (PCP) complexes, and three reconstituted light-harvesting systems, where an N-terminal construct of native PCP from Amphidinium carterae has been reconstituted with chlorophyll (Chl) mixtures: with Chl a, with Chl b and with both Chl a and Chl b. Using laser excitation into peridinin (Per) absorption band we take advantage of sub-picosecond energy transfer from Per to Chl that is order of magnitude faster than the Förster energy transfer between the Chl molecules to independently populate each Chl in the complex. The results indicate that reconstituted PCP complexes contain only two Chl molecules, so that they are spectroscopically equivalent to monomers of native-trimeric-PCP and do not aggregate further. Through removal of ensemble averaging we are able to observe for single reconstituted PCP complexes two clear steps in fluorescence intensity timetraces attributed to subsequent bleaching of the two Chl molecules. Importantly, the bleaching of the first Chl affects neither the energy nor the intensity of the emission of the second one. Since in strongly interacting systems Chl is a very efficient quencher of the fluorescence, this behavior implies that the two fluorescing Chls within a PCP monomer interact very weakly with each other which makes it possible to independently monitor the fluorescence of each individual chromophore in the complex. We apply this property, which distinguishes PCP from other light-harvesting systems, to measure the distribution of the energy splitting between two chemically identical Chl a molecules contained in the PCP monomer that reaches 280 cm(-1). In agreement with this interpretation, stepwise bleaching of fluorescence is also observed for native PCP complexes, which contain six Chls. Most PCP complexes reconstituted with both Chl a and Chl b show two emission lines, whose wavelengths correspond to the fluorescence of Chl a and Chl b. This is a clear proof that these two different chromophores are present in a single PCP monomer. Single molecule fluorescence studies of PCP complexes, both native and artificially reconstituted with chlorophyll mixtures, provide new and detailed information necessary to fully understand the energy transfer in this unique light-harvesting system.

Ensemble and single-molecule spectroscopy demonstrates that both emission and absorption of peridinin−chlorophyll−protein photosynthetic antennae can be largely enhanced through plasmonic interactions. We find up to 18-fold increase of the chlorophyll fluorescence for complexes placed near a silver metal layer. This enhancement, which leaves no measurable effects on the protein structure, is observed when exciting either chlorophyll or carotenoid and is attributed predominantly to an increase of the excitation rate in the antenna. The enhancement mechanism comes from plasmon-induced amplification of electromagnetic fields inside the complex. This result is an important step toward applying plasmonic nanostructures for controlling the optical response of complex biomolecules and improving the design and functioning of artificial light-harvesting systems.Strong enhancement of electromagnetic fields generated through plasmon resonances in metal films and particles has recently stimulated a considerable interest in diverse research fields such as optical spectroscopy, cell imaging, quantum information processing, nanophotonics, and biosensors. [1][2][3][4][5] This versatility results from a dramatic influence that plasmons impose on the absorption and emission properties of nearby located dipoles, for example, semiconductor nanocrystals and nanowires 6-12 or dye molecules. [13][14][15][16][17][18] Optical response of an emitter coupled to a plasmonic structure depends upon spatial arrangement as well as spectral characteristics of a studied system. Remarkable progress has been made in on-demand design of metal nanostructures, which is essential for tuning the resonance frequency and thus the coupling strength. 13,14,19 Complementary efforts focused on developing advanced experiments to study dipoles placed in the vicinity of a metal nanoparticle have shed light on the interplay between radiative and nonradiative processes in these systems. 16,18 This very relation determines whether the fluorescence is enhanced [9][10][11]16 or quenched due to the dominating role of nonradiative energy transfer from the dipole to the metal. 15,18 Metal-enhanced fluorescence (MEF) has been observed for many hybrid systems that include nanocrystals on corrugated metal surfaces, 10,11 dye molecules coupled to metal nanoparticles, 18 and nanocrystal-nanoparticle bioconjugates. 8In all these cases, very stable and highly fluorescing emitters have been selected. It would be, however, highly desirable to apply MEF to weakly fluorescing systems such as DNA, 20 carbon nanotubes 21 or, yet experimentally unexplored in this context, light-harvesting complexes. These latter proteinpigment systems, which contain chlorophyll (Chl) and carotenoid molecules embedded in a protein matrix, participate in the photosynthesis process by collecting sunlight energy and transferring it to reactions centers. The presence of fluorescing Chls and the protein, separated by a few nanometers, renders light-harvesting complexes ideal for studying the prote...

We combine ensemble and single-molecule spectroscopy to gain insight into the energy transfer between chlorophylls (Chls) in peridinin-chlorophyll-protein (PCP) complexes reconstituted with Chl a, Chl b, as well as both Chl a and Chl b. The main focus is the heterochlorophyllous system (Chl a/b-N-PCP), and reference information essential to interpret experimental observations is obtained from homochlorophyllous complexes. Energy transfer between Chls in Chl a/b-N-PCP takes place from Chl b to Chl a and also from Chl a to Chl b with comparable Förster energy transfer rates of 0.0324 and 0.0215 ps(-1), respectively. Monte Carlo simulations yield the ratio of 39:61 for the excitation distribution between Chl a and Chl b, which is larger than the equilibrium distribution of 34:66. An average Chl a/Chl b fluorescence intensity ratio of 66:34 is measured, however, for single Chl a/b-N-PCP complexes excited into the peridinin (Per) absorption. This difference is attributed to almost three times more efficient energy transfer from Per to Chl a than to Chl b. The results indicate also that due to bilateral energy transfer, the Chl system equilibrates only partially during the excited state lifetimes.

Reconstitution of the 16 kDa N-terminal domain of the peridinin-chlorophyll-protein, N-PCP, with mixtures of chlorophyll a (Chl a) and Chl b, resulted in 32 kDa complexes containing two pigment clusters, each bound to one N-PCP. Besides homo-chlorophyllous complexes, hetero-chlorophyllous ones were obtained that contain Chl a in one pigment cluster, and Chl b in the other. Binding of Chl b is stronger than that of the native pigment, Chl a. Energy transfer from Chl b to Chl a is efficient, but there are only weak interactions between the two pigments. Individual homo-and hetero-chlorophyllous complexes were investigated by single molecule spectroscopy using excitation into the peridinin absorption band and scanning of the Chl fluorescence, the latter show frequently well resolved emissions of the two pigments.