Cupredoxins

Figure 2. The cupredoxin fold in (a) the odd and (b) the even domains of hCP. The P-barrels comprise two sheets of P-strands. Sheet 1 involves strands 2a, 8, 7, and 4 and the four type I copper binding residues emanate from this sheet (see Table 3). Sheet 2 involves strands 2a, 1, 3, and 6.

Blue (or type 1 ) copper proteins (or cupredoxins) are important components of biological electron transfer processes in many organisms ranging from bacteria to animals, from fungi to plants.56 They are characterized by ... 

Another interesting blue protein is stellacyanin (FW = 20 000) from the Japanese lacquer tree Rhus vernicifera, in which, with respect to the other cupredoxins, glutamine replaces the methionine ligand.64 Stellacyanin also bears an overall positive charge (p/=9.9). It, therefore, gives a reversible Cu(II)/Cu(I) response at a glassy carbon electrode in aqueous solution (pH 7.6).61 The formal electrode potential of the Cu(II)/Cu(I) reduction (E01 = + 0.18 V vs. NHE) is the lowest among cupredoxins. 

A further cupredoxin we consider is mavicyanin from green zucchini peelings (FW = 12 700),70 a positively charged phytocyanin (p/= 8.85) the copper site of which is very similar to that of the above discussed... 

Finally, we examine azurin, a blue protein (FW = 14 000) devoted to bacterial electron transport, the copper centre of which has a penta-coordinate trigonal bipyramidal geometry, at variance with all the other cupredoxins, Figure 39.73... 

For example, in Chapter 12, Section 4, we have examined the electrochemical response of azurin (from Pseudomonas aeruginosa), the only cupredoxin in which the copper(II) ion is pentacoordinate. Its reversible Cu(II)/Cu(I) reduction occurs at Eol= +0.31 V, vs. NHE, at 25° C. Measurements of the variation of the formal electrode potential with temperature in a non-iso thermic electrochemical cell gives the two diagrams illustrated in Figure ll.20... 

The crystal structure of the pseudoazurin from Alcaligenes faecalis S-6 sometimes referred to as the blue protein (also as cupredoxin), has been reported to 2.0 A [74]. The protein folds in /3-sandwich which is described as being similar to plastocyanin and azurin. 

It has long been known that, under some conditions at least, electron transfer between the c and d hemes of the P aeruginosa enzyme is slow and requires times of the order of seconds (22). What does this mean It is not necessarily related to the loss of the hydroxide ligand from the d heme iron, because under some experimental conditions used, azurin (a cupredoxin) was present and the enzyme was reduced at the outset,... 

It is nearly 50 years since a c-type cytochrome was shown to catalyze peroxidase activity in crude extracts of Pseudomonas fluorescens (40). The enzyme responsible was first purified some 20 years later by Ell-folk and Soninen from the closely related P. aeruginosa and shown to be a diheme cytochrome c peroxidase (CCP) (41). These bacterial diheme CCPs are quite distinct from the superfamily of plant and yeast peroxidases (42) and are widely distributed among the Gram-negative bacteria (41, 43 6). Diheme CCPs are located in the periplasm (Fig. 2), where they catalyze the two-electron reduction of H2O2 to H2O by soluble one-electron donors such as cytochromes c and cupredoxins. 

II. Cupredoxins Proteins That Bind Only Type I Copper. 148... 

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... 

Spectra, but, in general, leaves the copper site the most exposed of the four cupredoxins. The sequence of Cbp is quite similar to that of stella-cyanin. Stellacyanin is a plant protein, also of unknown function, having visible spectra characteristic of type I copper, but lacking the methionine ligand found in all other type I proteins. A disulfide bond has been suggested as a potential copper ligand in stellacyanin the Cbp has both a methionine and the disulfide, so that prior to the structure determina-... 

A recently characterized single-domain copper protein, auracyanin (Trost et al., 1988), is a dimeric protein which has a visible spectrum more like that of the A. faecalis cupredoxin (pseudoazurin, subgroup II see Table II) than that of either azurin or plastocyanin, but, because of its cysteine content and rhombic EPR, it has been put in the other class in Table II. 

In summary, then, comparison of the four known cupredoxin structures helps reveal the essence" of this kind of protein. Schematically (see Fig. 6), it can be seen that the jS-barrel structure is composed of three parts an amino-terminal portion, a middle portion, and a carboxy-terminal portion (a functional grouping which probably is not relevant to the folding of the polypeptide). One part of the amino-terminal loop... 

The location of the copper with respect to the Greek key fold is interesting when compared to that of the cupredoxins. While the copper in the cupredoxins lies in the interior of the /8 barrel bound by three interior-facing residues of the carboxy-terminal loop in the )8 barrel, and by a histidine in an adjacent strand, the copper in SOD lies on the outside of its jS barrel, bound by one residue from the carhoxy-terminal loop and three from the adjacent strand (cf. Figs. 2c-5c with Fig. 8c.) A structural comparison of plastocyanin and SOD, coupled with sequence alignment of plastocyanin and ceruloplasmin (Ryden, 1988), showed that three of the SOD ligands correspond to putative copper ligands in ceruloplasmin. Why this is so will become more evident after the description of the ascorbate oxidase structure and its relationship to ceruloplasmin. 

The nature of the copper in these proteins is not totally clear. Dooley et al (1988) reported that the Achromobacter protein may have two kinds of type I sites in a total of three copper sites per dimeric protein, while the A. faecalis protein was reported to be a tetrameric protein with both type I and type II coppers (KakutanielaZ., 1981). Interestingly, the Achromobacter protein is green. Both of these nitrite reductases accept electrons from a cupredoxin. 

Cupredoxin fold , 47 95 Cupric fluoride, stereochemistry, 2 20 Cupric ion, exchange with cuprous ion, NMR of, 4 276-277... 

Tab. 5 Potential values for copper(ll/l) proteins (principally cupredoxins) in aqueous solution (all values are presumed to be for 25 °C, p, 0.1 M)...

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