Energy and Electron Transfer upon Selective Femtosecond Excitation of Pigments in Membranes of Heliobacillus mobilis Ursula Liebl, ‡,§ Jean-Christophe Lambry, ‡ Winfried Leibl, § Jacques Breton, § Jean-Louis Martin, ‡ and Marten H. Vos* ,‡ Laboratoire d’Optique Applique ´ e, INSERM U451, CNRS URA 1406, Ecole Polytechnique-ENSTA, 91120 Palaiseau, France, and SBE/DBCM, CEN de Saclay, 91191 Gif-sur-YVette Cedex, France ReceiVed February 26, 1996; ReVised Manuscript ReceiVed May 6, 1996 X ABSTRACT: Excitation energy transfer steps in membranes of Heliobacillus mobilis were directly monitored by transient absorption spectroscopy with a time resolution of 30 fs under selective excitation within the inhomogeneously broadened bacteriochlorophyll g Q Y band. The initial anisotropy was found to be >0.4, indicating that the pigments are excitonically coupled. After initial decay of this anisotropy in <50 fs, major sub-picosecond components associated with spectral equilibration were identified, corresponding to uphill energy transfer with a 300 fs time constant (812 nm excitation) and downhill energy transfer with 100 and 500 fs components (770 nm excitation). These equilibrations are ascribed predominantly to single excitation transfer steps, as anisotropy measurements showed that equilibration within spectrally similar pigments occurs on the same time scale as spectral equilibration, a situation which contrasts with that in photosystem I. Downhill energy transfer occurs to a significant extent directly to an energetically heterogeneous population of excited states as well as in a sequential way via gradually lower-lying pools of bacteriochlorophyll g. This finding supports a description in which all pigments, including the blue- most absorbing, are spatially organized in a random way rather than in clusters of spectrally similar species. Spectral equilibration is not entirely completed prior to formation of the primary radical pair P798 + A 0 - , which was found to proceed in a multiexponential way (time constants of 5 and 30 ps). No indication for the formation of radical species other than P798 + A 0 - on the time scale up to 100 ps was found. The bacteriochlorophyll pigments bound to photosynthetic proteins are involved in both energy transfer and primary photochemistry, processes which have an extremely high overall quantum efficiency. The bulk of these pigments serves to capture photons and to transfer the resulting excited state energy to the pigments constituting the primary donor, where photochemistry starts. As a general feature, the excited state for most pigments has a higher energy than the primary donor and for a minority of pigments it has a lower energy. Such an arrangement is thought to optimize the overall efficiency of photosynthesis (Trissl, 1993). In the photosynthetic membrane of purple bacteria and in photosystem II (PS II), 1 the two functions of energy and electron transfer, respectively, take place in distinct entities, the light-harvesting and reaction center complexes, and can therefore be studied separately. By contrast, in PS I and in the photosystems of heliobacteria and green sulfur bacteria, a pigment-protein complex exists in which both processes take place. Within this group of photosystems, the helio- bacteria are unique in that all energy transfer and primary electron transfer occurs in a single bacteriochlorophyll- containing protein complex, called the reaction center- antenna complex (RC) [for recent reviews, see Amesz (1995), Blankenship (1994), and van Grondelle et al. (1994)]. This complex binds a limited number of pigments (∼50, see below) and can be studied directly in membrane fragments. The most extensively studied representatives of the he- liobacteria, Heliobacterium chlorum (Gest & Favinger, 1983) and Heliobacillus mobilis (Beer-Romero & Gest, 1987) (the latter is used in the present study), have virtually identical spectroscopic properties (Beer-Romero et al., 1988). The RC core is a homodimer (Liebl et al., 1993), as is that of green sulfur bacteria (Bu ¨ttner et al., 1992), in contrast to the heterodimeric protein core of PS I and all other RCs known. The heliobacterial RC binds approximately 50 molecules of bacteriochlorophyll (Bchl) g, a type of Bchl found only in heliobacteria (Brockmann & Lipinski, 1983). Primary electron transfer occurs from the primary donor P798, a dimer of Bchl g [or Bchl g′, see van de Meent et al. (1990)], to the primary acceptor A 0 , identified as 8 1 - hydroxychlorophyll a (van de Meent et al., 1991). The overall process of energy transfer to P798 and charge separation has been reported to occur in 20-30 ps (Trost & Blankenship, 1989; van Noort et al., 1992; Lin et al., 1994a). Due to the spectral overlap of P798 with other Bchl g pigments, the intrinsic P798*A 0 f P798 + A 0 - charge separa- tion time (P798* denoting the singlet excited state of P798) cannot be measured independently. Using trap-limited energy transfer models, this time has been estimated to be less than 1.2 ps (Lin et al., 1994a), which is somewhat faster than the primary charge separation in wild type RCs of purple bacteria (Martin & Vos, 1992). The absorption spectrum of the membrane-bound RC complex has a dominant Bchl g Q Y band with a maximum ‡ INSERM U451. § SBE/DBCM. X Abstract published in AdVance ACS Abstracts, July 1, 1996. 1 Abbreviations: Bchl, bacteriochlorophyll; DAS, decay-associated spectrum; EDTA, ethylenediaminetetraacetic acid; FMO, Fenna- Matthews-Olson; fwhm, full width at half-maximum; LH1, light- harvesting complex 1; MOPS, 3-(N-morpholino)propanesulfonic acid; PMS, N-methyldibenzopyrazine methosulfate; PS, photosystem; RC, reaction center; SVD, singular value decomposition. 9925 Biochemistry 1996, 35, 9925-9934 S0006-2960(96)00462-X CCC: $12.00 © 1996 American Chemical Society