Fluorescence Decay Kinetics of Wild Type and D2-H117N Mutant Photosystem II Reaction Centers Isolated from Chlamydomonas reinhardtii Heather G. Johnston, † Jun Wang, ‡ Stuart V. Ruffle, ‡,§ Richard T. Sayre ,* ,‡ and Terry L. Gustafson* ,‡ Department of Chemistry, The Ohio State UniVersity, 100 West 18th AVenue, Columbus, Ohio 43210 and Department of Plant Biology, The Ohio State UniVersity, 1735 Neil AVenue, Columbus, Ohio 43210 ReceiVed: October 6, 1999; In Final Form: March 9, 2000 We compare the chlorophyll fluorescence decay kinetics of the wild type and the D2-H117N mutant photosystem II reaction centers isolated from Chlamydomonas reinhardtii. The histidine residue located at site 117 on the D2 polypeptide of photosystem II is a proposed binding site for one of two peripheral accessory chlorophylls located in the reaction center complex. The peripheral accessory chlorophylls are thought to be coupled with the primary electron donor, P680, and thus involved in energy transfer with P680. The conservative replacement of the histidine residue with an asparagine residue allows the chlorophyll to remain bound to the reaction center. However, slight changes in the structural organization of the reaction center may exist that can affect the energy transfer kinetics. We show that the D2-H117N mutation causes a shift in the 20-30 ps lifetime component that has been associated with energy equilibration among coupled chlorophylls in the photosystem II reaction center. Introduction Photosystem II is a protein-pigment complex present in the thylakoid membranes of higher plant and green alga chloroplasts that catalyzes the light-dependent oxidation of water. Chloro- phylls present in the PS II complex function in energy harvesting or as the primary electron donor during charge separation. Charge separation initiates electron transfer in the PS II reaction center resulting in the removal of electrons from water and release of molecular oxygen. 1 Since it was first isolated, the PS II reaction center has been determined to consist of the D1/ D2 polypeptides, the cytochrome b559 subunits, and the psb I protein. 2 Although the exact structure of the reaction center is still unknown, the D1/D2 polypeptides have been found to be highly homologous to the L/M polypeptides of the purple bacteria photosynthetic reaction center (PBPRC), whose struc- ture is known. 2-5 One difference existing between the two structures is the number of chlorophylls present. The PBPRC has four bound chlorophylls while the PS II reaction center contains approximately six chlorophylls/two pheophytin. 4,6 Based on analogies to the PBPRC two of the six chlorophylls may function as the primary electron donor, P680. 3,4,7 However, as many as six chlorins may be excitonically coupled. 8 Two accessory chlorophyll monomers lying in close proximity to the primary electron donor are believed to be involved in electron transfer to pheophytin, but also are coupled to P680. Two additional chlorophylls of the PS II reaction center, the peripheral accessory chlorophylls, are located further away from P680. 9 The peripheral accessory chlorophylls are thought to act as a link for energy transfer from the antennae chlorophylls of the PS II complex to P680. 1,10 The mechanism for energy transfer in the PS II reaction center is not well understood and remains an active area of research. Several models for energy transfer in the PS II reaction center have been put forth. 11-19 One current model by Holzwarth and co-workers is that after initial excitation there is a subpicosecond equilibration of energy among the primary donor and coupled chlorophylls in the reaction center complex. They assign a longer 20-40 ps lifetime to energy equilibration between the primary electron donor, additional excitonically coupled chlorophylls, most likely the peripheral accessory chlorophylls, and any additional chlorophylls bound to the reaction center. After energy equilibration the primary charge separation (P680 + / Pheo - ) then occurs in ∼3-8 ps. 11,13,20,21 Another interpretation of the energy transfer between the chlorophylls present in the reaction center has been put forth by Giorgi et al. 16,18 This model still assigns a subpicosecond energy equilibration among the primary electron donor and coupled chlorophylls. However, this model suggests that energy equilibration among all the chlo- rophylls present in the reaction center and the primary electron donor occurs on this subpicosecond time scale. After energy equilibration among the pigments they assign a 20-30 ps lifetime to the primary charge separation (P680 + /Pheo - ). Site-directed mutagensis has proven to be a valuable tool in gaining further insight into the functionality of the PS II reaction center. It is now possible to make specific changes in the amino acid sequence of the D1/D2 polypeptides. The kinetics of energy transfer between pigments in mutant PS II reaction centers can then be compared to the wild-type PS II reaction centers. Two sites of interest in the PS II reaction center are histidines located at site 118 on the D1 polypeptide and site 117 on the D2 polypeptide. 4 These histidines are proposed binding sites for the peripheral accessory chlorophylls. 22-24 A conservative replacement of the histidine residue would still allow the chlorophyll to be bound while possibly making changes in the distance and the orientation of the chlorophyll with respect to P680. A nonconservative replacement of the histidine residue * To whom correspondence should be addressed: sayre.2@osu.edu, gustafson.5@osu.edu. † Department of Chemistry. ‡ Department of Plant Biology. § Current Address: Department of Biomolecular Sciences, UMIST, Manchester, M60 1 QD, UK. 4777 J. Phys. Chem. B 2000, 104, 4777-4781 10.1021/jp993556l CCC: $19.00 © 2000 American Chemical Society Published on Web 05/11/2000