Azurin, domain structure

The subunits are arranged in the crystals as homotetramers with D2 symmetry. The structure of a subunit is shown schematically in Fig. 1 (87). Each subunit of 552 amino acid residues has a globular shape with dimensions of 49 x 53 x 65 A and is built up of three domains arranged sequentially on the polypeptide chain, tightly associated in space. The folding of all three domains is of a similar jS-barrel type. It is distantly related to the small blue copper proteins, for example, plastocyanin or azurin. Domain 1 is made up of two four-stranded jS-sheets (Fig. lb), which form a jS-sandwich structure. Domain 2 consists of a six-stranded and a five-stranded jS-sheet. Finally, domain 3 is built up of two five-stranded jS-sheets that form the jS-barrel structure and a four-stranded j8-sheet that is an extension at the N-terminal part of this domain. A topology diagram of ascorbate oxidase for all three domains and of the related structures of plastocyanin and azurin is shown in Fig. 2. Ascorbate oxidase contains seven helices. Domain 2 has a short a-helix (aj) between strands A2 and B2. Domain 3 exhibits five short a-helices that are located between strands D3 and E3 (a ), 13 and J3 (a ), and M3 and N3 (a ) as well as at the C terminus (ag and a ). Helix 2 connects domain 2 and domain 3. 

A comparison of the different variants of the jS-barrel domain structure in Fig. 1 shows that domain 1 of ascorbate oxidase has the simplest /3-barrel with only two four-stranded /8-sheets. Plastocyanin and azurin are quite similar but between strands 4 (El) and 6 (FI) they have insertions of one strand (plastocyanin) or one strand and an a-helix (azurin). Domain 2 has one additional strand H2 in sheet D next to strand E2 (sheet B and strand El in domain 1) and two additional strands, F2 and G2, in sheet C next to strand 12 (sheet A and strand FI in domain 1). Domain 3 resembles domain 2 except for the insertion of the short a-helices and the addition of the four-stranded /8-sheet at its N terminus. 

The first class is cupredoxins—single-domain blue copper proteins composed of only one BCB domain. These proteins include plastocyanin, azurin, pseudoazurin, amicyanin, auracyanins, rusticyanin, halocyanin, and sulfocyanin (see Section IV). Plantacyanin of the phytocyanin family (Section V), subunit II of the cytochrome c oxidase, and the recently characterized nitrosocyanin also fall into this class. The last two are single BCB domain polypeptides closely related structurally to cupredoxins, but harboring, respectively, a binuclear copper site known as CuA and a novel type of copper-binding site called red (see Sections IX and X). 

At this writing, the three-dimensional sttuctures of eight different naturally occurring type 1 copper proteins are known. These include the cupredoxins plastocyanin at 1.33 A resolution (pdb code 1 PTC), azurin at 1.8 A (pdb code 2AZA), pseudoazurin at 1.55 A (pdb code IPAZ), amicyanin at 1.3 A (pdb code lAAC), auracyanin at 1.55 A (pdb code IQHQ), rusticyanin at 1.9 A (pdb code IRCY), and the phytocyanins cucumber basic protein at 1.8 A (pdb code2CBP), and stellacyanin at 1.6 A (pdb code IJER) Atomic coordinates for these and all other single-domain type 1 copper proteins are available from the Research Collaboratory for Structural Bioinformatics (RCSB) Protein Data Bank (PDB) and can be accessed online at www.rcsb.org/pdb/. 

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