Kinetics of photo-induced electron transfer from high-potential iron-sulfur protein to the photosynthetic reaction center of the purple phototroph Rhodoferax fermentans.

Abstract

The kinetics of photo-induced electron transfer from high-potential iron-sulfur protein (HiPIP) to the photosynthetic reaction center (RC) of the purple phototroph Rhodoferaxfermentans were studied. The rapid photooxidation of heme c-556 belonging to RC is followed, in the presence of HiPIP, by a slower reduction having a second-order rate constant of4.8 x 107 M -l s -1. The limiting value of k.bs at high HiPIP concentration is 95 s-1. The amplitude of this slow process decreases with increasing HiPIP concentration. The amplitude of a faster phase, observed at 556 and 425 nm and involving heme c-556 reduction, increases proportionately. The rate constant of this fast phase, determined at 425 and 556 nm, is -3 x 105 s-1. This value is not dependent on HiPIP concentration, indicating that it is related to a first-order process. These observations are interpreted as evidence for the formation of a HiPIP-RC complex prior to the excitation flash, having a dissociation constant of -2.5 ,uM. The fast phase is absent at high ionic strength, indicating that the complex involves mainly electrostatic interactions. The ionic strength dependence of kobs for the slow phase yields a second-order rate constant at infinite ionic strength of 5.4 x 106 M-t.s-I and an electrostatic interaction energy of -2.1 kcal/mol (1 cal = 4.184 J). We conclude that Rhodoferax fermentans HiPIP is a very effective electron donor to the photosynthetic RC.Anoxygenic phototrophic bacteria contain two membranebound components that are essential for energy production, the photosynthetic reaction center (RC) and the cytochrome bc1 complex. Two types of RC have been structurally characterized: the RC from Rhodobacter sphaeroides is made of three subunits (1, 2), while, in addition to these, RC from Rhodopseudomonas viridis contains a fourth tetraheme cytochrome c subunit (2, 3). InRps. viridis the four heme groups have a bands at 559, 552, 556, and 554 nm, and reduction potentials of +380 mV, +20 mV, +310 mV, and -60 mV, respectively (4). The four hemes are arranged in an almost linear fashion, (2,5,6) and are aligned in the sequence P/c-559/c-552/c-556/c-554, where P indicates the bacteriochlorophyll special pair (4, 7). The highest potential heme (c-559) is oxidized the most rapidly (t/2 -300 ns) by the photooxidized P (P+), followed by a slower oxidation of heme c-556 (ti/2 2 ,us) (4,8,9). The quinol generated by RC photooxidation is utilized by the bc1 complex to reduce cytochrome c2 (10), a soluble protein closely related to mitochondrial cytochrome c (11,12). Cytochrome c2 then reduces the photooxidized RC, closing the photocycle. However, although cytochrome c2 reacts directly with P+ in Rb. sphaeroides (13-21), in Rps. viridis, it reacts with the tetraheme subunit (22-24). Only about half of the purple bacterial species described so far contain cytochrome c2 (25), and it is not known how the remaining species carry out electron transfer between the bc1 and RC complexes.The RC from the purple nonsulfur bacterium Rhodoferax fermentans (26)...

X-Ray Structure of Rhodobacter Capsulatus Cytochrome bc1: Comparison with its Mitochondrial and Chloroplast Counterparts

“…While the mitochondrial and bacterial enzymes are referred to as cyt bc 1 , the homologous enzyme in plant and algal chloroplasts and cyanobacteria is called plastohydroquinone : plastocyanin oxidoreductase or cyt b 6 f (for a review see Cramer et al 1996;Cramer et al 2004). In all cases, these energy transducing enzymes transfer electrons from a hydroquinone derivative (QH 2 ) (usually ubihydroquinone, menahydroquinone or plastohydroquinone), to an electron carrier molecue such as a c type cyt, a high potential iron sulfur protein (HiPIP) or a plastocyanin ( (Jenney and Daldal 1993;Jenney et al 1994;Hochkoeppler et al 1996;Kerfeld et al 1996) ). They are central players in respiration and photosynthesis as they contribute to the generation of an electrochemical potential ∆µH + , subsequently used for ATP production via the ATP synthase (Mitchell 1976;Dutton et al 1998;Saraste 1999).…”

“…These two proteins interact physically with very different electron acceptors: a c-type cyt (cyt c, c 2 , c y or c 8 ) (Jenney et al 1994;Kerfeld et al 1996) or a high potential iron-sulfur protein (HiPIP) (Hochkoeppler et al 1996) in the case of the cyt c 1 versus plastocyanin or a c-type cyt (cyt c 6 ) in the case of the cyt f (Cramer et al 1996). Moreover, the pH of the environment in which these subunits perform their specific functions is very different between the bacteria, mitochondria and chloroplasts.…”

The Primary Structure of Rhodoferax fermentans High‐Potential Iron‐Sulfur Protein, an Electron Donor to the Photosynthetic Reaction Center

“…The photo-induced electron transfer from Rf: fermentans Hipip to the photosynthetic RC involves the formation of a Hipip/RC complex (Kd approximately 2.5 pM) characterized by mainly electrostatic interactions, involving 5 -6 specific ionic pairs [13]. In an effort to establish the protein surface patch that is involved in protein-protein complex formation between the soluble Hipips and membrane-bound redox complexes such as bc, and RC, we have followed the approach previously applied to cytochromes c 1491 and the photoactive yellow protein [50].…”

“…This process follows multiphasic kinetics, with a fast phase occumng within a HipiplRC complex [13]. The high value of this electron transfer rate (t,,* = 2.2 ps) suggests a physiological role for Hipip in photocyclic electron transfer [13]. Recent evidence pointing to a physiological role for Hipips has also been established in Ru.…”

“…Experiments of light-induced oxidation of RC and membrane-bound c-type haem proteins have shown that RJ: fermentans Hipip is able to rapidly reduce the photo-oxidized photosynthetic RC through the reduction of the membranebound c-556 haem [7, 121. This process follows multiphasic kinetics, with a fast phase occumng within a HipiplRC complex [13]. The high value of this electron transfer rate (t,,* = 2.2 ps) suggests a physiological role for Hipip in photocyclic electron transfer [13].…”