Chloroplasts plastocyanin

Integration of PS II and PS I via cyt b6f in chloroplasts (Z scheme of photosynthesis). Pheo a represents pheophytin pQA and pQB represent phytoquinone A and B, respectively A, and At represent electron acceptor chlorophylls, respectively pc represents plastocyanin and Fd represents ferridoxin. 

Let us start our examination with the prototypical blue protein plastocyanin, found in the thylacoid membrane of chloroplasts, where it acts as an electron carrier in photosynthesis (see Figure 1). As Figure 30 illustrates, the active site of plastocyanin is formed of a Cu(II) ion (pseudo)tetrahedrally coordinated to two histidine nitrogen atoms and... 

Plastocyanin from parsley, a copper protein of the chloroplast involved in electron transport during photosynthesis, has been reported to have a fluorescence emission maximum at 315 nm on excitation at 275 nm at pH 7 6 (2°8) gjncc the protein does not contain tryptophan, but does have three tyrosines, and since the maximum wavelength shifts back to 304 nm on lowering the pH to below 2, the fluorescence was attributed to the emission of the phenolate anion in a low-polarity environment. From this, one would have to assume that all three tyrosines are ionized. A closer examination of the reported emission spectrum, however, indicates that two emission bands seem to be present. If a difference emission spectrum is estimated (spectrum at neutral pH minus that at pH 2 in Figure 5 of Ref. 207), a tyrosinate-like emission should be obtained. 

A transit peptide consisting of a hydrophobic 66 amino acid long peptide interspersed with positively charged residues has been identified and sequenced [52]. This is initially attached to the 99 amino acids of the mature plastocyanin, and is responsible for taking the plastocyanin across membranes into the thylakoid region of the chloroplast. 

Like Complex III of mitochondria, cytochrome b6f conveys electrons from a reduced quinone—a mobile, lipid-soluble carrier of two electrons (Q in mitochondria, PQb in chloroplasts)—to a water-soluble protein that carries one electron (cytochrome c in mitochondria, plastocyanin in chloroplasts). As in mitochondria, the function of this complex involves a Q cycle (Fig. 19-12) in which electrons pass, one at a time, from PQBH2 to cytochrome bs. This cycle results in the pumping of protons across the membrane in chloroplasts, the direction of proton movement is from the stromal compartment to the thylakoid lumen, up to four protons moving for each pair of electrons. The result is production of a proton gradient across the thylakoid membrane as electrons pass from PSII to PSI. Because the volume of the flattened thylakoid lumen is small, the influx of a small number of protons has a relatively large effect on lumenal pH. The measured difference in pH between the stroma (pH 8) and the thylakoid lumen (pH 5) represents a 1,000-fold difference in proton concentration—a powerful driving force for ATP synthesis. 

Cyanobacteria can synthesize ATP by oxidative phosphorylation or by photophosphorylation, although they have neither mitochondria nor chloroplasts. The enzymatic machinery for both processes is in a highly convoluted plasma membrane (see Fig. 1-6). Two protein components function in both processes (Fig. 19-55). The proton-pumping cytochrome b6f complex carries electrons from plastoquinone to cytochrome c6 in photosynthesis, and also carries electrons from ubiquinone to cytochrome c6 in oxidative phosphorylation—the role played by cytochrome bct in mitochondria. Cytochrome c6, homologous to mitochondrial cytochrome c, carries electrons from Complex III to Complex IV in cyanobacteria it can also carry electrons from the cytochrome b f complex to PSI—a role performed in plants by plastocyanin. We therefore see the functional homology between the cyanobacterial cytochrome b f complex and the mitochondrial cytochrome bc1 complex, and between cyanobacterial cytochrome c6 and plant plastocyanin. 

The extent to which an electron carrier is oxidized or reduced during photosynthetic electron transfer can sometimes be observed directly with a spectrophotometer. When chloroplasts are illuminated with 700 nm light, cytochrome/, plastocyanin, and plastoquinone are oxidized. When chloroplasts are illuminated with 680 nm light, however, these electron carriers are reduced. Explain. 

Figure 2-16 Beta cylinders. (A) Stereoscopic a-carbon plot of plastocyanin, a copper-containing electron-transferring protein of chloroplasts. The copper atom at the top is also shown coordinated by the nitrogen atoms of two histidine side chains. The side chains of the aromatic residues phenylalanines 19,29, 35,41, 70, and 82 and tyrosines 80 and 83 are also shown. Most of these form an internal cluster.

The simpler cytochrome bc] complexes of bacteria such as E. coli,102 Paracoccus dentrificans,116 and the photosynthetic Rhodobacter capsulatus117 all appear to function in a manner similar to that of the large mitochondrial complex. The bc] complex of Bacillus subtilis oxidizes reduced menaquinone (Fig. 15-24) rather than ubiquinol.118 In chloroplasts of green plants photochemically reduced plastoquinone is oxidized by a similar complex of cytochrome b, c-type cytochrome /, and a Rieske Fe-S protein.119 120a This cytochrome b6f complex delivers electrons to the copper protein plastocyanin (Fig. 23-18). 

Figure 23-17 The zigzag scheme (Z scheme) for a two-quantum per electron photoreduction system of chloroplasts. Abbreviations are P680 and P700, reaction center chlorophylls Ph, pheophytin acceptor of electrons from PSII QA, Qg, quinones bound to reaction center proteins PQ, plastoquinone (mobile pool) Cyt, cytochromes PC, plastocyanin A0 and Aj, early electron acceptors for PSI, possibly chlorophyll and quinone, respectively Fx, Fe2S2 center bound to reaction center proteins FA, FB, Fe4S4 centers Fd, soluble ferredoxin and DCMU, dichlorophenyldimethylurea. Note that the positions of P682, P700, Ph, Qa/ Qb/ Ay and A, on the E° scale are uncertain. The E° values for P682 and P700 should be for the (chlorophyll / chlorophyll cation radical) pair in the reaction center environment. These may be lower than are shown.

The electron donor to Chl+ in PSI of chloroplasts is the copper protein plastocyanin (Fig. 2-16). However, in some algae either plastocyanin or a cytochrome c can serve, depending upon the availability of copper or iron.345 Both QA and QB of PSI are phylloquinone in cyanobacteria but are plastoquinone-9 in chloroplasts. Mutant cyanobacteria, in which the pathway of phylloquinone synthesis is blocked, incorporate plasto-quinone-9 into the A-site.345a Plastoquinone has the structure shown in Fig. 15-24 with nine isoprenoid units in the side chain. Spinach chloroplasts also contain at least six other plastoquinones. Plastoquino-nes C, which are hydroxylated in side-chain positions, are widely distributed. In plastoquinones B these hydroxyl groups are acylated. Many other modifications exist including variations in the number of iso-prene units in the side chains.358 359 There are about five molecules of plastoquinone for each reaction center, and plastoquinones may serve as a kind of electron buffer between the two photosynthetic systems. 

ATP synthesis in chloroplasts. The flow of electrons between PSII and PSI (Fig. 23-18) is of great importance for ATP formation. As previously mentioned, plastocyanin is usually the immediate donor to P700 and serves as a mobile carrier to bring electrons to this reaction center. In this function it is analogous to cytochrome c of mitochondrial membranes. The essentiality of plastocyanin was shown by study of copper-deficient Scenedesmus (Fig. 1-11). The photoreduction of C02 by H2 is impaired in these cells, but the Hill reaction occurs at a normal rate. 

The plastocyanins are found in plant chloroplasts and other photosynthetic organisms, and act as membrane-bound electron carriers between photosystems II and I in the photosynthetic pathway of higher plants, green algae and some blue-green algae. 

Plastocyanin consists of a single polypeptide chain of molecular weight around 10 500 and one copper atom. It is synthesized in the cytoplasm as a precursor of higher molecular weight,911 with an additional polypeptide that is essential for transport of the protein into the chloroplast. Within... 

The plastocyanins are blue copper proteins found in the chloroplasts of higher plants and algae where they mediate electron transport between cytochrome f and P-700 (Barber, 1983 Haehnel, 1984, 1986 Cramer etal., 1985 Sykes, 1985 Andersen et al., 1987). Plastocyanins each contain one copper bound by a single polypeptide chain of molecular weight around 10500 (Sykes, 1985). The spectroscopic properties of the copper are those of a typical blue site. The properties of the plastocyanins have been the subject of detailed reviews (Sykes, 1985 Haehnel, 1986 Chapman, 1991). 

Figure 2. Schematic of photoinduced electron transport and phosphorylation reactions considered to occur in chloroplast lamellae [from Moreland and Hilton (2)]. Open arrows indicate light reactions solid arrows indicate dark reactions and the narrow dashed line represents the cyclic pathway. Abbreviations used PS I, photosystem I PS II, photosystem II Y, postulated electron donor for photosystem II Q, unknown primary electron acceptor for photosystem II PQ, plastoquinones cyt b, b-type cytochromes cyt f, cytochrome f PC, plastocyanin P700, reaction center chlorophyll of photosystem I FRS, ferredoxin-reducing substance Fd, ferredoxin Fp, ferredoxin-NADP oxidoreductase FeCy, ferricyanide asc, ascorbate and DPIP, 2,6-dichloropheno-lindophenol. The numbers la, lb, 2, 3, and 4 indicate postulated sites of action by...

Figure 5-19. Schematic representation of reactions occurring at the photosystems and certain electron transfer components, emphasizing the vectorial or unidirectional flows developed in the thylakoids of a chloroplast. Outwardly directed election movements occur in the two photosystems (PS I and PS II), where the election donors are on the inner side of the membrane and the election acceptors are on the outer side. Light-harvesting complexes (LHC) act as antennae for these photosystems. The plastoquinone pool (PQ) and the Cyt b(f complex occur in the membrane, whereas plastocyanin (PC) occurs on the lumen side and ferredoxin-NADP+ oxidoreductase (FNR), which catalyzes electron flow from ferredoxin (FD) to NADP+, occurs on the stromal side of the thylakoids. Protons (H+) are produced in the lumen by the oxidation of water and also are transported into the lumen accompanying electron (e ) movement along the electron transfer chain.

Complex III is also analogous to the Cyt bef complex of chloroplasts, both with respect to contents (two Cyt b s, one Cyt c, an Fe-S pro tern, and a quinone) and function within the membranes (e.g., interaction with a quinol) the isolated Cyt b f complex can also pass electrons to Cyt c as well as to its natural electron acceptor, plastocyanin. Complex III is also structurally and functionally analogous to a supramolecular protein complex in bacteria. 

Plastocyanins are the most widely studied cupredoxins. They are one of the most abundant copper proteins in plant photosynthetic tissues. Plant plastocyanins have an intricate evolutionary history because of their ancient bacterial origin. It is currently well accepted that plants diverged from the main eukaryotic domain into a separate lineage when the unicellular, oxygen respiring common ancestor of the eukaryotes incorporated a prokaryotic endosymbiont, the cyanobacterial chloroplast. 

Plant plastocyanins are synthesized in the cytosol as 160-170-ammo acid precursor polypeptides consisting of a 60-70-residue transit peptide followed by a 97 99-amino acid mature protein. The transit peptide imports the precursor plastocyanin molecule across the chloroplast envelope and thylakoid membranes to its final destination in the thylakoid lumen, where it shuttles electrons by accepting them from the membrane bound cytochrome / (cyt /) of the cyt b6/f complex and donating them to the photooxidized reaction center P700-I- of photosystem I. Cyanobacterial plastocyanins use an 30-amino acid leader seqnence for thylakoid membrane translocation. Currently, there are more than 100 plant and cyanobacterial plastocyanin sequences that are available either by direct protein sequencing or deduced from the nucleotide sequences of their genes. 

Cytochrome b f is used in place of cytochrome bc in cyanobacteria and chloroplasts. Light energy utilized by PSII results in plastoquinol production from plastoquinone. Reducing equivalents carried by the plastoquinol are then transferred across b(,f, through cytochromes bound to b(,f, to plastocyanin or cytochrome c. Reduced plastocyanin or cytochrome ce is then used to rereduce PSI. The crystal structure of cytochrome b(,f at 3.0 A resolution is shown... 

The photosynthetic apparatus of green plants and cyanobacteria oxidizes water and transfers electrons to NADP, with a net gain in electrochemical potential of 1.13 eV (at pH 7), utilizing the energy of two light quanta per electron. The complete system is contained in the chloroplasts, and is localized within the thylakoid membranes, with the exception of the electron carrier ferredoxin, which is in solution in the stroma, and serves to transfer electrons from the reducing end of photosystem I (PS I) to a membrane-bound flavoprotein which then reduces NADP, and of the copper protein plastocyanin (PC, the electron donor to PS I), which is in solution in the internal phase of thylakoids. 

If the model proposed by Andersson and Anderson [109] of total separation of PS I and PS II in the granal chloroplasts were to be accepted, electron transport from the PS II acceptors to P-700 would require a mobile electron carrier(s) which should diffuse laterally in the membrane fast enough to account for the observed electron transport rate. Plastoquinone [112] and plastocyanin are the candidates of choice for this role. The former has been shown to be present at approximately the same activity in the partitions and in the stroma-exposed membranes [43], while PC is known to be located in the intrathylakoid space [113],... 

Most of the thylakoid proteins are organized into four intrinsic protein complexes PS II complex, Cyt b/f complex, PS I complex and ATP synthetase (Fig. 1). The electron transport complexes are linked by mobile electron transport carriers, plastoquinone, plastocyanin and ferredoxin (see Chapter 10). Furthermore, chloroplasts that possess Chi b have the major light-harvesting Chi a/h-proteins of PS II (LHC II) that may represent over 50% of the thylakoid protein [13], as well... 

Hill reaction. In 1939, Robert Hill discovered that chloroplasts evolve O2 when they are illuminated in the presence of an artificial electron acceptor such as ferricyanide [Fe3+(CN)g]3-. Ferricyanide is reduced to ferrocyanide [Fe +(CN)5]4- in this process. No NADPH or reduced plastocyanin is produced. Propose a mechanism for the Hill reaction. 

The general function of this complex is that of transferring electrons from ubiquinone (or plastoquinone) to a hydrophilic protein acceptor (cytochrome c or plastocyanin). Therefore, in bacterial photosynthesis, it catalyzes the recycling of electrons from the secondary electron acceptor (Qn) to the secondary electron donor (cyt. Cj), completing thereby the cyclic electron transfer system. In chloroplasts and cyanobacteria, an analogous system transfers the electrons from plastoquinone (the secondary acceptor of PSII, A, 3) to plastocyanin (the secondary donor to PSI, 0, 2) and provides in this way an intersystem redox connection between PSII and PSI. The same complex is also involved in the cycling of electrons around PSI. 

The components of the quinol-cytochrome c (plastocyanin) oxidoreductase of chloroplasts, cyanobacteria and photosynthetic bacteria have been demonstrated to be very similar. This analogy proves the substantial unity of the mechanism of electron flow in all photosynthetic systems. For this reason the different components of the complexes will be discussed unitarily in the following sections in order to emphasize the functional and structural similarities between them. 

Cytochromes of b type are invariably involved in electron transfer in photosynthetic bacteria and in plant chloroplasts and cyanobacteria, and take active part in the mechanism of the quinol-cytochrome c (plastocyanin) oxidoreductase complex. 

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