Study of the high-potential iron sulfur protein in Halorhodospira halophila confirms that it is distinct from cytochrome c as electron carrier

Abstract

The role of high-potential iron sulfur protein (HiPIP) in donating electrons to the photosynthetic reaction center in the halophilic ␥-proteobacterium Halorhodospira halophila was studied by EPR and time-resolved optical spectroscopy. A tight complex between HiPIP and the reaction center was observed. The EPR spectrum of HiPIP in this complex was drastically different from that of the purified protein and provides an analytical tool for the detection and characterization of the complexed form in samples ranging from whole cells to partially purified protein. The bound HiPIP was identified as iso-HiPIP II. Its Em value at pH 7 in the form bound to the reaction center was Ϸ100 mV higher (؉140 ؎ 20 mV) than that of the purified protein. EPR on oriented samples showed HiPIP II to be bound in a well defined geometry, indicating the presence of specific protein-protein interactions at the docking site. At moderately reducing conditions, the bound HiPIP II donates electrons to the cytochrome subunit bound to the reaction center with a half-time of <11 s. This donation reaction was analyzed by using Marcus's outer-sphere electron-transfer theory and compared with those observed in other HiPIP-containing purple bacteria. The results indicate substantial differences between the HiPIP-and the cytochrome c 2-mediated re-reduction of the reaction center.electron transfer ͉ photosynthesis ͉ electron paramagnetic resonance ͉ redox potential ͉ reaction center H igh-potential iron sulfur proteins (HiPIPs) are soluble electron carriers containing a single cubane [Fe 4 S 4 ] cluster [for reviews, see refs. 1 and 2] found in both photosynthetic (3-6) and nonphotosynthetic (7, 8) proteobacteria, as well as in members of the so-called FBC group (flexibacter, bacteroides, cytophaga group) (9). Despite three decades of detailed biophysical and biochemical characterization, the role of HiPIPs as physiological electron donors to the reaction center (RC) in photosynthesis and to oxidase in respiration was not demonstrated until 1995 (10-12).HiPIPs are commonly regarded as exotic substitutes for the ''normal'' soluble carrier cytochrome c. This notion is based on the presence of soluble cytochromes in mitochondrial respiration, in cyanobacterial photosynthesis, and in a variety of other prokaryotic energy conserving systems. Moreover, the usual ''workhorses'' in the study of purple bacterial photosynthesis, Rhodobacter sphaeroides, Rhodobacter capsulatus, and Blastochloris viridis, do not contain HiPIP. Instead, cytochrome c 2 , together with its various iso-forms, functions in their bioenergetic chains. Several decades of studies on these species have resulted in an understanding of the structural and functional details of electron donation from cytochrome c 2 to the photosynthetic RC (see, for example, refs. 13-19). In contrast, the interaction between HiPIP and RC is still poorly understood. Except for the case of Rubrivivax gelatinosus (20-23), only scarce data are available (24, 25). Surveys of photosynthetic electron transfer among p...

Menaquinone as pool quinone in a purple bacterium

“…The redox midpoint potential of the P/P ϩ couple therefore is a key parameter indicative for the operating redox range of the whole chain. This E m value of P was determined by equilibrium optical redox titrations on isolated RCs and on membranes as well as through the redox potential dependence of the flash-induced absorption changes in membranes as described previously (13). All these experiments yielded the same result of E m7.5 ϭ ϩ270 Ϯ 10 mV [see Fig.…”

“…In H. halophila, this electron carrier is a high-potential iron sulfur protein (HiPIP) (13). The redox potential of this HiPIP [E m ϭ ϩ165 mV (13)] turned out to be substantially lower than that of other purple bacterial HiPIPs (14) and instead falls within the range of potentials generally observed for electron donors to RCs in phototrophs using MK as pool quinone (15). Because no purple bacterium operating its photosynthetic chain on MK has been described thus far, this finding is surprising.…”

Metalloproteins Containing Cytochrome, Iron–Sulfur, or Copper Redox Centers

“…872 However, other studies have shown a role for HiPIPs distinct from that of cytochrome c . 881 HiPIPs are also found in the membrane of some thermophilic organisms. 879 HiPIPs are mainly found in organisms with a photosynthetic reaction center having a tetraheme cytochrome (THC) subunit.…”

Nonredundant Roles for Cytochrome c 2 and Two High-Potential Iron-Sulfur Proteins in the Photoferrotroph Rhodopseudomonas palustris TIE-1

“…Many bacteria contain both small cytochromes and HiPIPs and sometimes contain multiple copies of each, a redundancy that has yet to be fully explained. Biophysical studies have shown that, in at least some of the organisms containing a HiPIP and a small cytochrome, both are capable of reducing the reaction center (5)(6)(7)(8). Under a given condition, a single protein (either the HiPIP or the cytochrome) generally appears to be the dominant electron donor (6).…”