Pathways of Energy Transformation in Antenna Reaction Center Complexes of Heliobacillus mobilis ² Sieglinde Neerken,* Thijs J. Aartsma, and Jan Amesz Department of Biophysics, Huygens Laboratory, Leiden UniVersity, P.O. Box 9504, 2300 RA Leiden, The Netherlands ReceiVed October 20, 1999; ReVised Manuscript ReceiVed December 28, 1999 ABSTRACT: The conversion of excitation energy in the antenna reaction center complex of Heliobacillus mobilis was investigated at 10 K as well as at 275 K by means of time-resolved absorbance difference spectroscopy of isolated membranes in the (sub)picosecond time range. Selective excitation of the primary electron acceptor, chlorophyll (Chl) a 670, and of the different spectral pools of bacteriochlorophyll (BChl) g (BChl g 778, BChl g 793, and BChl g 808) was applied. At 10 K, excitation at 770 or 793 nm resulted on the one hand in rapid energy transfer to BChl g 808 and on the other hand in fast charge separation from excited BChl g 793 (∼1 ps). Once the excitations were on BChl g 808, the bleaching band shifted gradually to the red, from 806 to 813 nm, and charge separation from excited BChl g 808 occurred by a very slow process (∼500 ps). The main purpose of our experiments was to answer the question whether an “alternative” pathway for charge separation exists upon excitation of Chl a 670. Our measurements showed that the amount of oxidized primary donor (P798 + ) relative to that of excited BChl g produced by excitation of Chl a 670 was considerably larger than upon direct excitation of BChl g. This indicates the existence of an alternative pathway for charge separation that does not involve excited antenna BChl g. This effect occurred at 10 K as well as at 275 K. The mechanism for this process is discussed in relation to different trapping models; it is concluded that charge separation occurs directly from excited Chl a 670. The traditional scheme for the primary processes in photosynthesis involves (i) the absorption of light by antenna pigments, (ii) transfer of excitation energy to the primary electron donor, and (iii) the transfer of an electron from the primary donor to an acceptor molecule. However, there are several aspects to this scheme that are the subject of discussion. First of all, it is not always clear whether step ii or step iii is rate-limiting in the generation of the charge-separated state, and both possibilities have, e.g., been considered for purple bacteria (1, 2). Second, the mechanism for so-called uphill transfer of excitation energy from the antenna to the reaction center, which appears to occur even at liquid helium temperature in heliobacteria and various species of purple bacteria (3, 4), is not understood. Finally, it has been proposed that alternative pathways may exist for charge separation that do not involve the excited state of the primary electron donor (4-7). Recently we performed time-resolved studies of the excited states and charge separation in reaction center core complexes of the green sulfur bacterium Prosthecochloris (Ptc.) aes- tuarii (7, 8). These complexes contain about 16 bacterio- chlorophylls (BChl) 1 a, two of which form the special pair P840, and four chlorophyll (Chl) a molecules absorbing near 670 nm (Chl a 670) (9, 10). Comparison of the population of excited BChl a and the extent of subsequent charge separation brought about by excitation either of BChl a or of Chl a 670 showed that upon excitation of the latter pigment an alternative pathway of charge separation existed. The phenomenon appeared to occur at 10 K as well as at 275 K. The question now arises whether a similar pathway might exist in heliobacteria. Like the green sulfur bacteria, the heliobacteria have a type I reaction center (11, 12). In contrast to other photosynthetic organisms they have a single pigment protein complex, which was called the ARC (antenna reaction center) complex (13) and which can be isolated after detergent solubilization (13, 14). The visible absorption spectrum of the isolated ARC complex is identical to that of whole cells and cytoplasmic membranes (13). The complex contains about 35 molecules of BChl g (15), a pigment related to BChl a, but with an ethylidene group at carbon C8 (ring II) (16). It absorbs in the Q y region with three bands peaking near 778, 793, and 808 nm, which are only resolved at low temperature. The corresponding spectral forms have been called BChl g 778, BChl g 793, and BChl g 808 (17). Only for BChl g 793 is there evidence for strong excitonic interaction (17). The primary electron donor, P798, is a dimer of BChl g, presumably of its 13 2 epimer (18). The ARC complex also contains two molecules of 8 1 - hydroxy Chl a (15). The latter pigment has a Q y absorption band at 668 nm and appears to act as primary electron acceptor, A 0 (19). Although chemically not identical, we shall call it Chl a 670, as in green sulfur bacteria (8). Nuijs et al. (19) were the first to apply pump-probe absorption spectroscopy to the study of energy transforma- ² This work was supported by the Section for Earth and Life Sciences (ALW) of The Netherlands Science Foundation (NWO). * Author to whom correspondence should be addressed: e-mail sigi@biophys.leidenuniv.nl; fax: +31 71 5275819. 1 Abbreviations: A0, primary electron acceptor; ARC, antenna reaction center; BChl, bacteriochlorophyll; Chl, chlorophyll; P798, primary electron donor. 3297 Biochemistry 2000, 39, 3297-3303 10.1021/bi992433o CCC: $19.00 © 2000 American Chemical Society Published on Web 03/03/2000