Cucumber basic blue protein

There are a number of excellent sources of information on copper proteins notable among them is the three-volume series Copper Proteins and Copper Enzymes (Lontie, 1984). A review of the state of structural knowledge in 1985 (Adman, 1985) included only the small blue copper proteins. A brief review of extended X-ray absorption fine structure (EXAFS) work on some of these proteins appeared in 1987 (Hasnain and Garner, 1987). A number of new structures have been solved by X-ray diffraction, and the structures of azurin and plastocyanin have been extended to higher resolution. The new structures include two additional type I proteins (pseudoazurin and cucumber basic blue protein), the type III copper protein hemocyanin, and the multi-copper blue oxidase ascorbate oxidase. Results are now available on a copper-containing nitrite reductase and galactose oxidase. 

Cucumber basic blue protein has been put in a class of its own in view of having a disulfide, unUke plastocyanin, although its fold is generally like that of plastocyanin. 

Cucumber basic blue protein (Cbp) is a protein without known function, also known as cusacyanin or plantacyanin. Its structure (Guss et al., 1988) completes the repertoire of cupredoxins with known structures. The topology of its folding is similar (Fig. 5) to those of plastocyanin and azurin, as might have been expected from sequence similarities and... 

Fig. 5. (a) Copper site in cucumber basic blue protein (Cbp). (b) Ribbon drawing of the Cbp backbone, (c and d) Schematic of Cbp topology. 

The cucumber basic blue protein is made up of a single polypeptide chain of 96 amino acids containing a single copper ion with a molecular weight of 10000 (Adman, 1985). The amino acid sequence (Bergman et al., 1977) is very similar to those of stellacyanin and umecyanin. The copper in cucumber blue protein displays the characteristic spectroscopic properties of a typical blue copper site (Table 5-5). 

The crystal structure of cucumber basic blue protein has now been refined to 3.0 A resolution (Adman, 1985). The protein consists of eight strands, only five of which form a P-sandwich and the protein has less P-sheet character than plastocyanin or azurin. The ligands to copper are provided by the side chains of His-39, Cys-79, His-84 and Met-89. The copper site has the N2SS coordination seen in plastocyanin. The imidazole rings of the His-39 and His-94 residues are exposed to the solvent providing a likely entry site for electon transfer to the copper centre. 

The type I copper sites function as electron transfer centers in the blue copper proteins and in multicopper enzymes, particularly oxidases (33). They are characterized by their intense blue color, their unusually small A values, and their very positive redox potentials (Table II). X-ray crystal structures of several blue copper proteins have been determined, notably plastocyanin (34), azurin (35), cucumber basic blue protein (36), and pseudoazurin (37). The active site structures show marked similarities but also distinct differences (Fig. 8). 

Basic blue proteins have been isolated from a number of plant sources and have previously been referred to as plantacyanin (Aikazyan and Nalbandyan, 1975,1979 Sakurai et al, 1982). The protein from cucumber has been the most extensively studied of the basic blue proteins and the crystal structure is now available (Guss et al., 1988). The function of this basic blue protein is unknown, however, it is probably not involved in photosynthetic electron transport as it will not replace plastocyanin in that electron transport chain (Adman, 1985). 

Unknown stellacyanin, umecyanin, cucumber basic blue copper protein... 

Bond Cu(SCPh3)L Azurin Populus plastocyanin Enteromorpha prolifera plastocyanin Pseudoazurin Cucumber basic blue copper protein"... 

The physiological functions of the phytocyanins are also currently unknown. This family of proteins consists of stellacyanin (20 kD) [97, 101], umecyanin (14 kD), the basic blue protein (from cucumber = plantacyanin) (8 kD), and cusacyanin (molecular masses see [17]). 

Upon perturbation of the geometry of the blue copper center, from plastocya-nin to cucumber basic blue to nitrite reductase (a green protein), flie Cys S n —>Cu CT intensity decreases while the Cys S cr—>Cu CT intensity increases... 

The molecular structure of another blue protein, the phytocyanin (phytocyanins are electron carriers found in the non-photosynthetic part of plants) cucumber basic protein (FW=10 100), also known as plantacyanin, is shown in Figure 33.60... 

Guss, J. M., et al. (1988). Phase determination by multiple-wavelength x-ray diffraction crystal structure of a basic blue copper protein from cucumbers. Science 241, 806-811. 

Blue copper proteins, 36 323, 377-378, see also Azurin Plastocyanin active site protonations, 36 396-398 charge, 36 398-401 classification, 36 378-379 comparison with rubredoxin, 36 404 coordinated amino acid spacing, 36 399 cucumber basic protein, 36 390 electron transfer routes, 36 403-404 electron transport, 36 378 EXAFS studies, 36 390-391 functional role, 36 382-383 occurrence, 36 379-382 properties, 36 380 pseudoazurin, 36 389-390 reduction potentials, 36 393-396 self-exchange rate constants, 36 401-403 UV-VIS spectra, 36 391-393 Blue species... 

We have studied the blue copper proteins plastocyanin, azurin, cucumber basic protein ( ) and nitrite reductase (NiR) (159,160). We shah focus on these four in the remainder of this section. 

Solvent is usually excluded from the blue copper site, which is buried 6 A inside the protein, having only the His ligand from the copperbinding loop exposed to the surface. The phytocyanins, stellacyanin and plantacyanin (cucumber basic protein), are exceptions, in which both His ligands are solvent exposed and the copper ion is only 3 A beneath the protein surface. This situation makes the copper center in this family of blue copper proteins more accessible to low-molecular-weight solutes (see Section V). 

Crystal structures of three phytocyanins are currently available. Two are for plantacyanins, from cucumber (also known as cucumber basic protein) (Guss et al., 1988, 1996) and from spinach (Einsle et al., 2000), and one is for the recombinant BCB domain of cucumber stella-cyanin (Hart et al., 1996). The three proteins display folding topology identical to one another, suggesting that phytocyanins fold into a uniform structure, which can be designated as a phytocyanin fold. As a historical note, the crystallization of the cucumber basic protein and its preliminary crystallographic data were reported in 1977, before any structure of a blue copper protein was available (Colman et al., 1977). However, the structure was solved in 1988 only by application of the then newly... 

Redox potentials for the different copper centers in the blue oxidases have been determined for all members of the group but in each case only for a limited number of species. The available data are summarized in Table VI 120, 121). The redox potentials for the type-1 copper of tree laccase and ascorbate oxidase are in the range of 330-400 mV and comparable to the values determined for the small blue copper proteins plastocyanin, azurin, and cucumber basic protein (for redox potentials of small blue copper proteins, see the review of Sykes 122)). The high potential for the fungal Polyporus laccase is probably due to a leucine or phenylalanine residue at the fourth coordination position, which has been observed in the amino-acid sequences of fungal laccases from other species (see Table IV and Section V.B). Two different redox potentials for the type-1 copper were observed for human ceruloplasmin 105). The 490-mV potential can be assigned to the two type-1 copper sites with methionine ligand and the 580-mV potential to the type-1 center with the isosteric leucine at this position (see Section V.B). The... 

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). 

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