Rusticyanin 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... 

In rusticyanin and pseudoazurin, the axial Met ligand adopts a different orientation than in azurin, resulting in a stronger Cu(II)-S8 Met bond and a tetragonal distortion. The magnetic anisotropy tensor is rhombic in the Co(II)-substituted proteins, and the pseudocontact contribution to... 

Rusticyanin has a high reduction potential (680 mV), which is similar to that for the Type 1 Cu center in fungal as opposed to tree laccase (785 mV) (73). This trend is so far unexplained. From the sequence and EXAFS studies, His-Cys-His-Met coordination is a reasonable possibility for rusticyanin (55). It may well be that the reduction potential is determined by effects of a polypeptide backbone on Cu—S(Cys) and Cu—S(Met) bond distances and the Cu ligand field (74). If this is the case, however, rusticyanin would be expected to have one or both Cu—S distances shorter than in other blue copper proteins, which is not borne out by information from EXAFS (Table IV). A further possibility that the Cu(I) form is three-coordinate, as in the case of plasto-cyanin at low pH (Fig. 2), has no strong support at present (75). 

Rusticyanin is an abundant, highly stable periplasmic blue copper protein (e.g., Cobley and Haddock 1975 Jedlicki et al. 1986 Ronk et al. 1991 Nunzi et al. 1993 Blake et al. 1993). Studies of rusticyanin include kinetic competence in the iron oxidation reaction (Blake and Shute 1994), identification of a His ligand to the copper center (Casimiro et al. 1995), a solution NMR structure (Botuyan et al. 1996), and a high-resolution X-ray structure (Walter et al. 1996). Rusticyanin forms a complex with new c-type heme cytochrome in iht A. ferrooxidans electron transport chain (Giudici-Orticoni et al. 2000). 

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. 16 a, b. Folding pattern of the small blue protein rusticyanin. The protein consists of thirteen /1-strands a ribbon model - the copper ion is represented by the blue circle [112] b disposition of ligands and spatially close side-chains in the copper site of one of the solution structures of Cu(I) rusticyanin. [112a] with permission... 

The conserved hydrogen bond network in cupredoxins is believed to play a role in maintaining the geometry of the copper center and in determining the relative constraints of the Cu and Cir sites, which in turn influences the relative stability of the two states of the protein and, ultimately, the reduction potential. " Indeed, mutations of the Asn in the extra-loop H-bond (i.e., the Asn next to the N-terminal His ligand) in both azurin and plastocyanin resulted in reduced stability and, in the case of azurin, an increase of the reduction potential from 286 mV for wild-type azurin to 396 mV for the variant protein. In rusticyanin, the Asn residue in the extra-loop H-bond is replaced with a Ser residue.Mutation of this Ser in rusticyanin resulted in reduced stability and a decrease of reduction potential by 110 mV. Based on these results, the authors... 

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