Poplar plastocyanin

A solution structure of French Bean plastocyanin has been reported by Wright and co-workers,19 using nuclear magnetic resonance techniques described in Section 3.5 of Chapter 3. The structure, determined from a plastocyanin molecule in solution rather than in a solid-state crystal, agrees well with that of reduced poplar plastocyanin X-ray crystallographic structure reported above. Conformations of protein side chains constituting the hydrophobic core of the French bean plastocyanin are well-defined by the NMR technique. Surface side chains show... 

Figure 30 X-Ray structure of the active site of poplar plastocyanin...

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...

Fig. 5. The a-carbon chain of poplar plastocyanin residues 50-64 from the X-ray crystal structure thin line), compared with that of S. obliquus plastocyanin obtained from a 2DNMR study [37a], showing the effect of deletions of residues 57 and 58...

The blue, or type 1, copper proteins, azurin from Pseudomonas aeruginosa (Adman et ai, 1978 Adman and Jensen, 1981) and from Al-caligenes denitrificans (Norris et al., 1983, 1986) and poplar plastocyanin (Guss and Freeman, 1983 Guss et al., 1986), have been studied by X-ray diffraction. These involve a Cu(I)/Cu(II) redox system. Cu(I) d ) is... 

Additional information has been obtained from single crystal, polarized optical and ESR spectroscopic studies924 on poplar plastocyanin, which have allowed a correlation of the electronic structure of the blue copper active site with its geometric structure. In summary, the three dominant absorption bands at 13 350, 16 490 and 17 870 cm-1 were assigned to CysS- Cu (d 2-,2 charge-transfer transitions. The methionine makes only a small contribution, due to the long Cu—S(Met) bond (2.9 A) and the poor overlap of the methionine sulfur orbitals with the dx y orbital of copper. Histidine-Cu charge transfer contributes to the weaker absorptions at 21 390 and... 

Fig. 5-9. The copper site in poplar plastocyanin based on the structure determined by Guss and Freeman (1983). Some of the bond angles and all of the copper-ligand bond distances are indicated.

Fig. 5-11. The effect of pH at the Cu(I) site in poplar plastocyanin as determined by Guss et al. (1986). Bond distances are shown in Angstroms and residue numbers are indicated in parentheses.

Figure 3. Blue copper proteins. A X-ray structure of poplar plastocyanin (21). B Absorption spectrum of plastocyanin and normal D 4 cCuCl42 (e scale expanded by 10). C X-band EPR spectrum of plastocyanin and Y>4 cCuCl42. ...

Figure 4. Single-crystal EPR of poplar plastocyanin (29) orientation of the dxz.y2 orbital. A Unit cell and molecular orientation with respect to the applied magnetic field. B EPR spectra and simulations for the crystal orientations shown. C Orientation of the g direction and the dx2.yz orbital superimposed on the blue copper site.

Figure 6. X-ray absorption spectroscopy. A Energy level diagram depicting a Cu Is - HOMO transition at 8979 eV. B Orientation averaged XAS spectra. C Polarized single-crystal XAS spectra for poplar plastocyanin. (Data are from reference 35.)...

Guss JM, Freeman HC (1983) Structure of oxidized poplar plastocyanin at 1.6 A resolution. J Mol Biol 169 521-563... 

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). 

Figure 6 Copper sites in (a) poplar plastocyanin, (b) A. den. azurin, (c) tomato plantacyanin, and (d) cucumber stellacyanin. Distances are for the oxidized form, with those for the reduced form in parentheses...

Oxidation state Poplar plastocyanin P. vulgaris plastocyanin S. oleracea plastocyanin P. aer. azurin ... 

The mononuclear copper site is located in domain 3 and has the four canonical type-1 copper ligands (His, Cys, His, and Met) also found in plastocyanin and azurin. It is coordinated to the NDl atoms of His 445 and 512, the SG atom of the Cys 507, and the SD atom of Met 517 in a distorted trigonal pyramidal geometry. The SD atom is at the long apex (see Fig. 4). Bond lengths of the type-1 copper for both subunits are displayed in Table III. They are compared with oxidized poplar plastocyanin (95) and azurin from Pseudomonas aeruginosa (96). Figure 5 shows an overlay of the type-1 copper site in azurin, plastocyanin, and ascorbate oxidase. 

Fig. 5. Stereo plot of the overlay of the type-1 copper sites for poplar plastocyanin (thin line), Pseudomonas aeruginosa azurin (medium line), and ascorbate oxidase (thick line).

The first crystal structure information on a blue copper protein, for poplar plastocyanin in the Cu(II) state, was published in 1978 (2, 3). Since then, the Cu(I) state and related apo and Hg(II) substituted forms (5, 6), the green algal plastocyanin from Enteromorpha prolifera [Cu(II)] (7), azurin from Alcaligenes denitrificans [Cu(II) and Cu(D] (8, 9), azurin from Pseudomonas aeruginosa [Cu(II)] (10, 11), as well as pseudoazurin from Alcaligenes faecalis S-6 (12), and the cucumber basic protein, both in the Cu(II) state, have been published (13), making this one of the best-documented class of proteins. In addition, information as to three-dimensional structure in solution has been obtained from two-dimensional NMR studies on French bean and Scenedesmus obliquus plastocyanins (14,15). This review is concerned in the main with the active site chemistry. Other recent reviews are listed (16-20). 

Bond Distances and Angles for the Cu Active Site of Poplar Plastocyanin"... 

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