Amicyanin ligands

Several copper enzymes will be discussed in detail in subsequent sections of this chapter. Information about major classes of copper enzymes, most of which will not be discussed, is collected in Table 5.1 as adapted from Chapter 14 of reference 49. Table 1 of reference 4 describes additional copper proteins such as the blue copper electron transfer proteins stellacyanin, amicyanin, auracyanin, rusticyanin, and so on. Nitrite reductase contains both normal and blue copper enzymes and facilitates the important biological reaction NO) — NO. Solomon s Chemical Reviews article4 contains extensive information on ligand field theory in relation to ground-state electronic properties of copper complexes and the application of... 

Fig. 6. Electron transfer complex between methylamine dehydrogenase and amicyanin from Paracoccus dentrificans (PDB Accession Code 2MTA). The distance shown is between eN of the redox cofactor tryptophan tryptophylquinone of methylamine dehydrogenase and the eN of the His-95 ligand of amicyanin.

Amicyanin is a type I copper protein in which two histidines, one cysteine and one methionine provide the four ligands for the redox-active copper. The P. denitrificans protein is composed of 105 amino acid residues. Its primary sequence is known (Van Spanning et al., 1990) and its crystal stmcture has been determined (Durley et al., 1993 Cunane et al., 1996). 

The redox properties of amicyanin are dependent on pH and also are affected by its association with MADH (Gray et al., 1988 Zhu et al., 1998). The pH dependence of the EmValue of amicyanin free in solution correlates with that of a single protonated ligand having a pK of 7.5. The crystal stmctures of oxidized amicyanin at pH 4.8 and reduced amicyanin at pH 4.4 show that the Hiscopper ligand is doubly protonated in the reduced... 

For electron transfer from copper to heme, two dominant sets of pathways of comparable efficiency were predicted. In each set of pathways, the point of intermolecular electron transfer was from the backbone O of Glu i of amicyanin to the backbone N of Gly of cytochrome c-551i, and the entry of electrons to iron occurred either via the porphyrin ring or the His ligand. In one set of pathways the exit of electrons from copper occurred via the Cys copper ligand, and the phenolic side chain of Tyr was an intermediate between Cys and Glu. In the other set of pathways the exit of electrons from copper occurred via the Met copper ligand, and the backbone of Lys was an intermediate between Met and Glu i... 

Figure 12 Hydrophobic patch surrounding the exposed edge of the His ligand, His 95, in amicyanin. (Reprinted with permission from Ref. 115. 1992 American Chemical Society)...

Binding of a paramagnetic, redox-inactive [Cr(CN)6]3- anion to specific sites of a blue copper protein, amicyanin, has been used in NMR-spectroscopic studies of the protein structure in solutions.285Ab initio calculations of the ligand-field spectra of [Cr(CN)6]3 have been performed and the results compared with those for cyano complexes of the other first-row transition metals.286 The role of Cr—C—N bending vibrations in the phosphorescence spectra... 

Table 5. Position of the copper ligands in plastocyanin, pseudoazurin, and amicyanin...

Fig. 14. Comparison of the amino acid sequences of plastocyanin (pcy), amicyanin (acy), and pseudoazurin (paz). The copper ligands are framed conserved residues are underlined. Based on Guss and Freeman 1983 [22], Ryden and Hunt 1993 [71], and Durley et al. 1994 [78]...

Rusticyanin s type 1 copper center with its ligands His 85, Cys 138, His 143, and Met 148 resembles those of the other small blue proteins. Although its tertiary structure is a /1-meander, there are distinct differences between the amino acid folding patterns of rusticyanin and the other small blue proteins. Plastocyanin contains 8 [22, 71], amicyanin 9 [20, 78], and rusticyanin 13 / -strands, respectively [112] (Fig. 16). 

Fig. 5. Representation of the Cu site of P. versutus amicyanin, highlighting the loop that contains the three copper ligands Cys-93, His-96, and Met-99. The copper atom is indicated by the dark sphere in the center of the figure.

As mentioned earlier in this section, three of the four active site ligands in cupredoxins are situated close together on a single loop (see Fig. 5). In Table III the loops found in this area of three different cupredoxins are shown. To investigate the role the length and composition of this loop have on the active site properties of amicyanin the corresponding loops from plastocyanin (113) and azurin (130) have been introduced. The properties of these mutants are compared to wild-type protein in Table IV. In both cases a novel site that is produced does not resemble, spectroscopically, wild-type amicyanin or... 

C-Terminal Ligand-Containing Loop of Paracoccus uersutus Amicyanin, Poplar Plastocyanin, Alcaligenes faecalis S-6 Pseudoazurin, and Pseudomonas aeruginosa Azurin... 

Before reviewing the details of the reactions between proteins from P. denitrificans it is important to first discuss the available structural information. The proteins MADH and amicyanin are known to associate quite strongly in solution (288). A binary complex has been crystallized and the structure (92) of part of the interface between the two proteins is shown in Fig. 20. The hydrophobic patch of amicyanin, which surrounds the exposed imidazole ring of the ligand His-96, is found associating with a mainly hydrophobic region on the L subunit of MADH. There are also interactions between amicyanin and the larger H subunit of MADH. The two proteins interact in such a way that the TTQ cofactor of MADH and the copper of amicyanin are approximately 9 A apart. 

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