Bacteriopheophytin

One of the most complex separation schemes utilizes flash liquid chromatography and PLC to obtain petropophyrins both from geochemical samples or those synthesized and used subsequently as standards [110]. Ocampo and Repeta [111] described the scheme of petroporphyrins isolation in which at the first step the sediment extract is fractionated into ten fractions on silica gel using dichlo-romethane (fractions 1 to 4), a mixture of dichloromethane-acetone with increasing acetone concentrations (for fractions 5 to 9), and, at last, dichlo-romethane methanol (4 1) (fraction 10). Next, the fifth fraction was separated on silica PLC plates using dichloromethane-acetone (97.5 2.5 v v v) as a developer. Two purple bands (with Rj 0.53 and 0.50) were recovered from silica and purified further on a silica gel column with dichloromethane-acetone (97.5 2.5, v v v) as an eluent. The emiched fraction was then separated by PLC with the same solvent mixture, and the purple bands containing two bacteriopheophytin allomers were recovered with acetone. 

For convenience of discussion, a schematic diagram of bacterial photosynthetic RC is shown in Fig. 1 [29]. Conventionally, P is used to represent the special pair, which consists of two bacterial chlorophylls separated by 3 A, and B and H are used to denote the bacteriochlorophyll and bacteriopheophytin, respectively. The RC is embedded in a protein environment that comprise L and M branches. The initial electron transfer (ET) usually occurs from P to Hl along the L branch in 1—4 picoseconds (ps) and exhibits the inverse temperature dependence that is, the lower the temperature, the faster the ET. It should be noted that the distance between P and Hl is about 15 A [53-55]. 

Once the special pair has absorbed a photon of solar energy, the excited electron is rapidly removed from the vicinity of the reaction centre to prevent any back reactions. The path it takes is as follows within 3 ps (3 X 10 12 s) it has passed to the bacteriopheophytin (a chlorophyll molecule that has two protons instead of Mg2+ at its centre), without apparently becoming closely associated with the nearby accessory bacteriochlorophyll molecule. Some 200 ps later it is transferred to the quinone. Within the next 100 ps the special pair has been reduced (by electrons coming from an electron transport chain that terminates with the cytochrome situated just above it), eliminating the positive charge, while the excited electron migrates to a second quinone molecule. 

There is rapid ( ps) electron transfer to a bacteriopheophytin molecule (BP). 

There is charge separation by electron transfer from bacteriopheophytin to a quinine (QA) and then on to a second quinine (QB) ... 

Photosystem 1 Reaction Center 4 bacteriochlorophylls, 2 bacteriopheophytins and a non-heme ferrous center [Fe(N-His) (0-Glu)] 131... 

Further work using time-resolved EPR and magnetophotoselection (MPS), using plane-polarized light to excite the triplet state, gave information on the orientation of the optical transition dipole axes relative to the principal axes of the triplet state. By this technique the transition moments of the primary donor"6, the carotenoid in the bRC"7 and the bacteriopheophytin in the inactive B branch 4>0"8 were determined. 

V). The centers resemble PSII of chloroplasts and have a high midpoint electrode potential E° of 0.46 V. The initial electron acceptor is the Mg2+-free bacteriopheophytin (see Fig. 23-20) whose midpoint potential is -0.7 V. Electrons flow from reduced bacteriopheophytin to menaquinone or ubiquinone or both via a cytochrome bct complex, similar to that of mitochondria, then back to the reaction center P870. This is primarily a cyclic process coupled to ATP synthesis. Needed reducing equivalents can be formed by ATP-driven reverse electron transport involving electrons removed from succinate. Similarly, the purple sulfur bacteria can use electrons from H2S. 

Reaction centers of purple bacteria. The exact composition varies, but the properties of reaction centers from several genera of purple bacteria are similar. In Rhodopseudomonas viridis there are three peptide chains designated H, M, and L (for heavy, medium and light) with molecular masses of 33,28, and 24 kDa, respectively. Together with a 38-kDa tetraheme cytochrome (which is absent from isolated reaction centers of other species) they form a 1 1 1 1 complex. This constitutes reaction center P870. The three-dimensional structure of this entire complex has been determined to 0.23-nm resolution288 319 323 (Fig. 23-31). In addition to the 1182 amino acid residues there are four molecules of bacteriochlorophyll (BChl), two of bacteriopheophytin (BPh), a molecule of menaquinone-9, an atom of nonheme iron, and four molecules of heme in the c type cytochrome. In 1984, when the structure was determined by Deisenhofer and Michel, this was the largest and most complex object whose atomic structure had been described. It was also one of the first known structures for a membrane protein. The accomplishment spurred an enormous rush of new photosynthesis research, only a tiny fraction of which can be mentioned here. 

Figure 23-31 (A) Stereoscopic ribbon drawing of the photosynthetic reaction center proteins of Rhodopseudomonas viridis. Bound chromophores are drawn as wire models. The H subunit is at the bottom the L and M subunits are in the center. The upper globule is the cytochrome c. The view is toward the flat side of the L, M module with the L subunit toward the observer. (B) Stereo view of only the bound chromophores. The four heme groups Hel-He4, the bacteriochlorophylls (Bchl) and bacteriopheophytins (BPh), the quinones QA and QB/ and iron (Fe) are shown. The four hemes of the cytochrome are not shown in...

These bacteria cannot in general oxidize water and must live on more readily oxidizable substrates such as hydrogen sulfide. The reaction centre for photosynthesis is a vesicle of some 600 A diameter, called the chromato-phore . This vesicle contains a protein of molecular weight around 70 kDa, four molecules of bacteriochlorophyll and two molecules of bacteriopheophy-tin (replacing the central Mg2+ atom by two H+ atoms), an atom Fe2+ in the form of ferrocytochrome, plus two quinones as electron acceptors, one of which may also be associated with an Fe2+. Two of the bacteriochlorophylls form a dimer which acts as the energy trap (this is similar to excimer formation). A molecule of bacteriopheophytin acts as the primary electron acceptor, then the electron is handed over in turn to the two quinones while the positive hole migrates to the ferrocytochrome, as shown in Figure 5.7. The detailed description of this simple photosynthetic system by means of X-ray diffraction has been a landmark in this field in recent years. 

Figure 5.7 Electron transfer processes in the first stages of photosynthesis. The energy of light E absorbed by the antenna chlorophylls is transferred to the special pair (BChl)FC is the ferrocytochrome, BPh the bacteriopheophytin and QFe, Q are quinones...

In Purple Bacterial Reaction Centers, Electrons Move from P870 to Bacteriopheophytin and Then to Quinones... 

Chlorophylls a and b contain the alcohol phytol bacteriochlorophylls a and b have either phytol or geranylger-aniol, depending on the species of bacteria. Photosynthetic organisms also contain small amounts of pheophytins or bacteriopheophytins, which are the same as the corresponding chlorophylls or bacteriochlorophylls except that two hydrogens replace the Mg2+ (see fig. 15.4). We will see that... 

Reaction centers of purple bacteria typically contain three polypeptides, four molecules of bacteriochlorophyll, two bacteriopheophytins, two quinones, and one nonheme iron atom. In some bacterial species, both quinones are ubiquinone. In others, one of the quinones is menaquinone (vitamin K2), a naphthoquinone that resembles ubiquinone in having a long side chain (fig. 15.10). Reaction centers of some species, such as Rhodopseudomonas viridis, also have a cytochrome subunit with four c-type hemes. 

The crystal structure of reaction centers from R. viridis was determined by Hartmut Michel, Johann Deisenhofer, Robert Huber, and their colleagues in 1984. This was the first high-resolution crystal structure to be obtained for an integral membrane protein. Reaction centers from another species, Rhodobacter sphaeroides, subsequently proved to have a similar structure. In both species, the bacteriochlorophyll and bacteriopheophytin, the iron atom and the quinones are all on two of the polypeptides, which are folded into a series of a helices that pass back and forth across the cell membrane (fig. 15.1 la). The third polypeptide resides largely on the cytoplasmic side of the membrane, but it also has one transmembrane a helix. The cytochrome subunit of the reaction center in R. viridis sits on the external (periplasmic) surface of the membrane. 

When P870 is excited, it transfers an electron to a bacteriopheophytin. The bacteriopheophytin then re-... 

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