Plastocyanin reaction sites

Metalloproteins fall into three main structure categories depending on whether the active site consists of a single coordinated metal atom, a metal-porphyrin unit, or metal atoms in a cluster arrangement. In the context of electron-transfer metalloproteins, the blue Cu proteins, cytochromes, and ferre-doxins respectively are examples of these different structure types. Attention will be confined here mainly to a discussion of the reactivity of the blue Cu protein plastocyanin. Reactions of cytochrome c are also considered, with brief mention of the [2Fe-2S] ferredoxin, and high potential Fe/S protein [HIPIP]. 

Earlier suggestions that the two uncoordinated and invariant residues His35 (inaccessible to solvent and covered by polypeptide) and His83 (remote and 13 A from Cu) are, from effects of [H ] on rate constants (and related pKg values), sites for electron transfer may require some re-examination. Thus, it has been demonstrated in plastocyanin studies [50] that a surface protonation can influence the reduction potential at the active site, in which case its effect is transmitted to all reaction sites. In other words, an effect of protonation on rate constants need not necessarily imply that the reaction occurs at the site of protonation. His35 is thought to be involved in pH-dependent transitions between active and inactive forms of reduced azurin [53]. The proximity of... 

Redox inert complexes [Cr(phen)3] and [CitCN) ] have been shown by nmr to bind at different points on the surface of the blue copper protein plastocyanin. Both sites are close to electron channels to the copper center and are the likely sites occupied by the oxidants [Co(phen)3] and [Fe(CN)6], which have been shown to bind to the protein.The reaction of [Co(phen)3] in inhibited by [Cr(phen)3]. ... 

While there is at present no full understanding as to why plastocyanin should require two sites for reaction, there is now much evidence detailing this two-site reactivity. Moreover, the recent X-ray crystal structure of ascorbate oxidase (which has 4 Cu atoms per molecule) has indicated a plastocyanin-like domain, with the two type 3 Cu s (in close proximity with the type 2 Cu) located at the remote site. Fig. 2 [5]. Since electrons are transferred, from the type 1 Cu to O2 bound at the type 3 center this structure defines two very similar through-bond routes for biological electron transfer. 

Table 2. Acid dissociation pK values, 1 = 0.10 M(NaCl), relating to the active site protonation of different plastocyanins, PCu(I), as determined by (a) proton NMR (b) the variation of rate constants (25 °C) with pH for the [FelCN) ] oxidation of PCu(I), 1 = 0.10 M(NaCl), and (c) similar experiments with [Co(phen)3] " as oxidant. The latter is an apparent value only, and is believed to be composite due to reaction occurring at the remote site...

For reaction to occur at the remote site of plastocyanin suggests that there is a preponderance of positively charged residues around the exposed heme edge of... 

Four products were obtained containing between 2 and 6 moles of amine per plastocyanin. More recently the reaction has been carried out to yield single and double modified products. No effect of modification at 42-45 on the rate of the PC u(I) reduction of P700 is reported, suggesting that these residues are not involved in the reaction. In a further elaboration the reaction of plastocyanin, cross-linked (by carbodiimide treatment) to cytochrome f at the remote site, with P700 has been studied and the effects of Mg " and Na" explored [158]. 

Parsley, spinach, French bean, poplar and S. obliquus (but not A. variabilis) conform extensively to the above criteria for reaction at the remote site. There is extensive evidence for cytochrome f reacting at the remote site on plastocyanin. The aromatic residue at 83 would seem to be a prime candidate as lead-in group for electron transfer. Desolvation at the surface around 83, and interaction with an aromatic component on the reaction partner, e.g. the porphyrin ring of cytochrome f, may be important. The exact manner of electron transfer has yet to be confirmed. The distance from the aromatic ring of Tyr83 to the Cu for electron transfer is 12 A. 

Cryokinetic studies of the plastocyanin-ferricyanide redox reactions in 50 50 v/v MeOH + H2O, pH = 7.0, p = 0.1 M reveal an Eyring plot shown for the second-order rate constant k from 25 °C to -35°C. The reaction is irreversible over the whole temperature range and there is no evidence for a change in the Cu(I) active site. Recalling that these reactions may involve consecutive steps, explain the deviation from a linear Eyring plot. F. A. Armstrong, P. C. Driscoll, H. G. Ellul, S. E. Jackson and A. M. Lannon, J. Chem. Soc. Chem. Communs. 234 (1988). 

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. 

The other way to study the "conductivity of protein molecules towards electron tunneling is to investigate the quenching of luminescence of electron-excited simple molecules by redox sites of proteins [95,96]. Experiments of this sort on reduced blue copper proteins have involved electron-excited Ru(II)(bpy)3, Cr(III)(phen)3, and Co(III)(phen)3 as oxidants. The kinetics of these reactions exhibit saturation at protein concentrations of 10 3 M, suggesting that, at high protein concentrations, the excited reagent is bound to reduced protein in an electron transfer precursor complex. Extensive data have been obtained for the reaction of reduced bean plastocyanin Pl(Cu(I)) with Cr(III)(phen)3. To analyze quenching experimental data, a mechanistic model that includes both 1 1 and 2 1 [Pl(Cu(I))/ Cr(III)(phen)3] complexes was considered [96]... 

Pinacolone, o-(diphenylphosphino)benzoyl-coordination chemistry, 401 Piperidine, IV-hydroxy-metal complexes, 797 pA a values azole ligands, 77 Plant roots amino acids, 962 carboxylic acids, 962 Plastocyanin copper binding site, 557 copper(II) complexes, 772 copper(II) site in, 770 Platinum, dichlorobis(benzonitrile)-IR spectrum, 264 Platinum, cis-dichlorodianunine-antitumor activity, 34, 979 Platinum, ethylenebis(triphenylphosphine)-reactions with 5,6-dimethyl-2,l,3-benzothiadiazole, 194 Platinum blue formation, 265 Platinum complexes acetylacetone reactions, 380 amides, 491 amidines... 

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...

In this complex, there are two optically active sites. Spinach plastocyanin is a type I copper protein, in which two reactive sites have been identified on its surface, at least. The electron transfer reaction occurs with significantly large stereoselectivity the ratio of the observed reaction rate constant (k /k ) is 1.6 to 2.0. The difference in the activation enthalpy, AAH a, is 3.0 kJ mol-1, and the difference in the activation entropy, AS (a-a) is 15 J mol-1 K-1. This means that the stereoselectivity arises from the entropy term. 

Crystallographic analysis has provided us with a detailed structure of hCp on the other hand, essentially all of the structure-function analyses have been done on FetSp. Also, except for the copper site structural homology, the two proteins are quite different. hCp is composed of six plastocyanin-like domains (plastocyanin is a type 1 copper-containing protein) that are arranged in a trigonal array (Zaitseva et al., 1996). One result of this domain replication is a conformational fold that produces a distinct, negatively charged patch on the protein surface adjacent to the catalytically active type 1 Cu(II). This copper atom is in domain 6. (Domains 2 and 4 contain type 1-like copper sites that do not participate in the ferroxidase reaction.) Lindley et al. (1997) have proposed that this... 

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