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J. Phys. Chem. 1996, 100, 3320-3322
Ultrafast Energy Transfer in Chlorosomes from the Green Photosynthetic Bacterium Chloroflexus aurantiacus Sergei Savikhin,† Yinwen Zhu,‡ Robert E. Blankenship,‡ and Walter S. Struve*,† Ames Laboratory and Department of Chemistry, Iowa State UniVersity, Ames, Iowa 50011, and Department of Chemistry and Biochemistry and Center for the Study of Early EVents in Photosynthesis, Arizona State UniVersity, Tempe, Arizona 85287-1604 ReceiVed: December 14, 1995X
Energy transfers between the bacteriochlorophyll c and a antennae in light-harvesting chlorosomes from the green bacterium Chloroflexus aurantiacus have been studied in two-color pump-probe experiments with improved sensitivity and wavelength versatility. The BChl c f BChl a energy transfers are well simulated with biexponential kinetics, with lifetimes of 2-3 and 11 ps. They do not exhibit an appreciable subpicosecond component. In the context of a kinetic model for chlorosomes, these lifetimes suggest that both internal BChl c processes and the BChl c f BChl a energy-transfer step contribute materially to the empirical rodto-baseplate energy-transfer kinetics.
Introduction Chlorosomes from the green photosynthetic bacterium Chloroflexus aurantiacus are light-harvesting bodies that contain some 104 bacteriochlorophyll (BChl) c pigments encased in an ellipsoidal lipid envelope.1-3 Electronic excitations in the 740 nm BChl c antenna of a chlorosome are transferred through a much smaller, lower-energy 795 nm BChl a-protein baseplate antenna (∼500 pigments) that interfaces the chlorosome to the B808-866 BChl a complexes and the reaction centers.4 The BChl c antenna is unique among photosynthetic light-harvesting systems in that it consists of BChl c oligomers, whose organization is determined by pigment-pigment rather than pigment-protein interactions.5 These oligomers form rodlike light-harvesting elements that are large enough to be visible under electron microscopy.1 Despite a number of fluorescence and pump-probe studies of the downhill BChl c f BChl a energy-transfer step in chlorosomes, its dynamics remain poorly understood. There is consensus that a major component of this energy transfer occurs with ∼10 ps kinetics in isolated chlorosomes (∼15 ps in whole cells), because several groups have observed 10-15 ps BChl c excitation decay components.5 Some of these laboratories have found concomitant BChl a excitation rise features with 10-15 ps kinetics as well. When examined under higher time resolution,6 the empirical BChl c decay kinetics exhibited additional major components with shorter lifetimes (1-2 ps and ∼100 fs). In two-color pump-probe experiments on chlorosomes from Cf. aurantiacus, we recently searched for evidence of equilibration between different BChl c spectral forms and investigated the details of BChl c f BChl a energy transfer.6 No photobleaching/stimulated (PB/SE) rise features were found for pump and probe wavelength combinations within the BChl c antenna Qy absorption spectrum, even though such features are readily distinguished with femtosecond kinetics in BChl a-protein antennae such as FMO trimers from the green sulfur bacterium Chlorobium tepidum7 and LH2 complexes from the purple bacterium Rhodobacter sphaeroides (Savikhin, S.; Struve, W., unpublished work). Hence, our experiments showed no evidence for equilibration between BChl c spectral forms, †
Iowa State University. Arizona State University. X Abstract published in AdVance ACS Abstracts, February 15, 1996. ‡
0022-3654/96/20100-3320$12.00/0
suggesting (in agreement with spectral hole-burning studies of BChl c antennae in whole cells of Cf. aurantiacus8) that the ∼20 nm fwhm BChl c Qy absorption band arises principally from homogeneous broadening. In two-color studies of the BChl c f BChl a energy transfer, no PB/SE rise features appeared when the pump and probe wavelengths were 760 and 800 nm, even though these wavelengths were situated within the BChl c and BChl a absorption regions, respectively. For the pump and probe wavelengths 790 and 820 nm, PB/SE rise components were found with lifetimes ∼100 fs, 2 ps, and 10 ps. These lifetimes mirrored the PB/SE decay components found using pump and probe wavelengths in the BChl c region (740-770 nm). These results led us to propose a kinetic model in which BChl c excitations equilibrate rapidly (,1 ps) with a BChl spectral form absorbing near 790 nm; subsequent energy transfers occurred to a longer-wavelength (∼795-800 nm) BChl a species with ∼10 ps kinetics.6 It was unclear whether the intermediate 790 nm BChl form (provisionally termed BChl a1 in ref 6) was a BChl c or a species, and there was no independent biochemical evidence for its existence. Recent technical improvements have enabled us to reinvestigate the energy-transfer kinetics in chlorosomes from Cf. aurantiacus. While our two-color technique offers jitter-free absorption difference profiles with high S/N, it was previously limited to pump and probe wavelengths that were separated by e30 nm. Further stabilization of our self-mode-locked Ti: sapphire laser output spectrum now permits the use of pumpprobe wavelength separations up to 60 nm. Our single-sideband radio-frequency multiple-modulation scheme has been superseded by a system that can detect absorption differences under excitation of one out of every ∼30 chlorosomes.9 The present work yields a different scenario for the BChl c f BChl a energy-transfer kinetics, probably because singlet-triplet annihilation in the BChl a antenna and the large SE cross section for the strongly coupled BChl c antenna combined to mask part of the BChl a rise kinetics for some probe wavelengths in the earlier work. Since the BChl a antenna contains some 20-fold fewer pigments than the BChl c antenna, singlet-triplet annihilation (due to sharp funneling of excitations into the baseplate antenna) can greatly reduce the observed BChl a absorption difference signal. In chlorosomes from green sulfur bacteria, the relative BChl a content is typically lower by a factor of ∼5 than in Cf. aurantiacus, with the result that van Noort et © 1996 American Chemical Society
Letters
J. Phys. Chem., Vol. 100, No. 9, 1996 3321
Figure 2. Kinetic scheme for energy transfers in chlorosomes from Cf. aurantiacus.
Figure 1. Absorption difference profiles for (a) 751 f 790 nm and (b) 751 f 810 nm two-color experiments on intact chlorosomes from Cf. aurantiacus. Negative-going signals are dominated by PB/SE at all times. Insets show the same profiles in 80 ps windows. Smooth curves give optimized fits to the profiles in 10 ps windows; fitting parameters are given in text.
al.10 were unable to determine the kinetics of BChl c/d f BChl a energy transfers in Chlorobium Vibrioforme. Materials and Methods Chlorosomes from Cf. aurantiacus were isolated according to the method of Gerola and Olson.11 The self-mode-locked Ti:sapphire laser and pump-probe apparatus were described previously. The radio-frequency (rf) multiple-modulation detection system was replaced by a new design, in which the probe beam detector photodiode was incorporated in an RLC prefiltering loop tuned to the rf detection frequency.9 Chlorosome samples were circulated through the laser beam at 4 m/s in a centrifugal sample cell, yielding a sample turnover time of ∼6 µs. A fresh chlorosome sample was used in each run. The average laser power in both beams combined was
