Plastocyanin spinach

Figure 3. Dependence of first-order rate constants ko6, (25 °C) vs. [Co(phen)s3+] for the oxidation of plastocyanin PCu(I). Conditions pH, 7.5 (phos) and I, 0.10 M (NaCl). Key , spinach and A, parsley. (Reproduced from Ref. 10. Copyright 1978, American Chemical Society.)...

Figure 5 of Valentine, J. S. DeFreitas, D. M. J. Chem. Ed., 1985, 62(11), 990-997. Copyright 1985, Division of Chemical Education, Inc.) (B) EPR spectrum for spinach plastocyanin. (Reprinted with permission from Figure 3b of Solomon, Edward I. Baldwin, Michael J. Lowery, Michael D. Chem. Rev., 1992, 92, 521-542. Copyright 1992, American Chemical Society.)... 

Figure 31 Cyclic voltammograms recorded at an edge-oriented pyrolitic graphite electrode in an aqueous solution (pH 6) of spinach plastocyanin. In the absence (a) and in the presence (b) of [Pt(NH3)6]4+. Scan rate 0.02 V s J...

Fig. 1. The a-carbon chain structure of poplar plastocyanin [16] including details of the active site and the remote acidic patches 42-45 and 59-61. The first of these has been modified to include an acidic residue at position 45 as e.g. for spinach and French bean plastocyanins...

The solution conformation of plastocyanin from French bean, spinach, and S. obliquus has now been determined from distance and dihedral angle constraints derived by NMR spectroscopy [37,40]. These two-dimensional NMR studies have indicated a well defined backbone conformation, which is very similar to that of poplar PCu in the crystalline state. However, in the case of S. obliquus there are deletions at positions S7 and 58 which influence the shape in the acidic region and in particular close to residues 59-61. The gap which is created is in effect repaired with consequent tightening of the loop 57-62 as indicated in Fig. 5. One of the pronounced bulges at the remote site of poplar and presumably other higher plant plastocyanins is not therefore present in S. obliquus (or plastocyanin from other green algae) [31, 32], as well as parsley... 

Table 4. Comparison of rate constants (25 °C) for the reactions of parsley and spinach plastocyanins, pH7.5,1=0.10 M(NaCl) [95, 99, 100]...

The kinetics of the reduction of spinach plastocyanin PCu(II) by the optically active complexes 2,6-bis[3-(S)- or 3-(/ )-carboxyl-2-azabutyl] pyridine, here abbreviated to (S,S)- or (f ,/ )-ALAMP have been studied [107]. The latter enantiomer (A-configuration) reacts 1.6-2.0 times faster at different values of pH and temperature than the S,S form. Activation parameters have shown that the observed stereoselectivity is a consequence of the difference in activation... 

Table 7. Partitioning of electron transfer between adjacent (k ) and remote (ke) binding sites on spinach plastocyanin PCu(I) at 25°C, pH7.5, I=0.10M(NaCl), using redox inactive [(NH3)5CoNHjCo(NH3)5]5+ [100, 117]...

The pKj values obtained for PCu(II) and PCu(I) are summarized in Table 9. Before discussing what appears to be a clear cut trend, the precision of individual pKa s should first be considered. It has for example been necessary to revise (downwards) the spinach PCu(II) pKa from 5.3 to 4.8 [121]. Plastocyanin is known to denature at or around pH 4.0 and in earlier work the lowest pH used was 4.5, which does not allow as accurate a fit to pK, values. By using a pH-jump method in which the final low pH is attained at the time of stopped-flow mixing (one reactant solution carrying substantially more buffer is allowed to control the pH), it is now possible with some confidence to include data down to pH 4.0. The other uncertainty is in the precision of the pKg for the PCu(I) remote site from a two pK fit. Again the data has to be free from artifacts introduced at... 

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. 

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. 

Fig. 5.39. Schematic drawing of the active site of P. aeruginosa azurin, spinach plastocyanin and cucumber stellacyanin (H = histidine, C = cysteine, M = methionine, G = glycine, Q = glutamine) (adapted from [198]).

Fig. 5.40. (A) H NMR spectra at 298 K of oxidized spinach plastocyanin at 800 MHz (adapted from [117]). (B) Far downfield region of the H NMR spectra of oxidized (i) P. aeruginosa azurin, (ii) spinach plastocyanin and (iii) cucumber stellacyanin containing signals not observable in direct detection (adapted from [198]). The positions and line widths of the signals were obtained using saturation transfer experiments by plotting the intensity of the respective exchange connectivities with the reduced species as a function of the decoupler irradiation frequency.

A small rhombic splitting in7the g values is also calculated (g g = 0.017). As a rhombic splitting was also indicated by our crystal field computations (11), a higher resolution EPR spectrum than the previously considered X-band data was obtained. Figure 18 shows the results of Q-band EPR on a frozen sample of spinach plastocyanin. A rhombic splitting of 0.017 is clearly discernable, in agreement with the adjusted Xa calculation. 

Figure 18. Q-band EPR spectrum of the g perpendicular region of spinach plastocyanin. Field sweep was between 11,275 and 12,275 gauss while the microwave frequency was 34.282 GHz.

Research aimed at identifying the ligands comprising the flattened tetrahedral blue copper center has been particularly intense in the case of plastocyanin. Direct evidence for a sulfur ligand has come from x-ray photoelectron spectral (XPS) experiments on bean plastocyanin, where a large shift of the S2p core energy of the single cysteine (Cys-85) residue in the protein upon metal incorporation (164.5, apo 169.8, native 168.8 eV, Co(II) derivative) was observed (15). The two histidines in spinach plastocyanin exhibit pK values below 5 in NMR titration experiments,... 

The first observation was reported in the electron transfer reaction between spinach plastocyanin with optically active iron(II) complexes, [Fe(S,S)-alamp] (A-form) and its (R,R)-isomer (A-form), where alamp is 2,6-bis[3-(S)- or 3-(R)-carboxy-2-azabutylpyridine (see Scheme 25) [56]. 

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. 

The stereoselective reduction of spinach plastocyanin with several cobalt cage complexes (Scheme 26) has been reported, too [60]. These cage complexes are very useful for investigation of outer-sphere electron transfer reactions because of their inertness to hydrolysis and to loss of ligands in the redox reaction. 

Reversible protonation and dissociation of the exposed His ligand have been observed in several BCP in the reduced metal-bound state. Since this protonation renders the proteins inactive, it has been characterized thoroughly (Sykes, 1985, 1991). An active site of 4.9 was determined by NMR for Cu(I) spinach plastocyanin (Markley et al., 1975). The occurrence of this process was conhrmed later by the crystal structure of reduced poplar plastocyanin at low pH (Cuss et al., 1986). Similar equilibria have been characterized in Achromobacter cycloclastes pseudoa-zurin (pA a 4.6) (Dennison et al., 1994b) and in Thiobacillus versutus ami-cyanin (pA a 6.7) (Lommen et al., 1988). In the latter system a lineshape analysis revealed that this His residue, on protonation and detachment from the copper(I) ion, fluctuates between two conformers (Lommen and Canters, 1990). 

PCF Cd II) plastocyanin complexed with cytochrome/) Spinacia oleracea (spinach) Ubbink and Bendall, 1997... 

In contrast, spinach plastocyanin binds to the soluble domain of its physiological partner cytochrome/ in a single orientation, indicating a short electron transfer path between the metal ions (Ubbink et al., 1998). A low-resolution structural model of the plastocyanin-cytochrome/complex was obtained by including paramagnetic constraints (derived from H and chemical shift differences) in molecular dynamics simulations where the structures of the two partners were kept rigid (Ubbink et al.,... 

The first NMR studies on the paramagnetic state of Ps. aeruginosa azurin (Hill et al., 1976) and spinach plastocyanin (Beattie et al., 1975 Markley et al., 1975) were reported together with experiments on the... 

Fig.3. 800-MHz NMR spectra of oxidized (A) Pseudomonas aeruginosa azurin, (B) spinach plastocyanin, and (C) cucumber stellacyanin recorded in D2O solution. The letters identify the resonance of the equivalent proton in the three proteins. In the insets the far-downheld regions containing signals not observable in direct detection are shown (Bertini et al., 2000). The positions and the linewidths of the signals of the oxidized species were obtained using saturation transfer experiments over the far-downheld region by measuring the intensity of the exchange connectivity with the corresponding signal in the reduced species (Bertini et al., 1999, 2000).

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