Redox properties of an engineered purple Cu(A) azurin.
Purple Cu(A) centers are a class of binuclear, mixed-valence copper complexes found in cytochrome c oxidase and nitrous oxide reductase. An engineered Cu(A) protein was formed by replacing a portion of the amino acid sequence that contains three of the ligands to the native type I copper center of Pseudomonas aeruginosa azurin with the corresponding portion of sequence from the Cu(A) center of cytochrome c oxidase from Paracoccus denitrificans [Proc. Natl. Acad. Sci. USA 93 (1996) 461]. Oxidation-reduction midpoint potential (E(m)) values of the Cu(A) azurin of +399+/-10 and +380+/-2mV, respectively, were determined by cyclic voltammetry and spectrochemical titration. An n value of one was obtained, indicating that the redox reaction is cycling between the mixed valence and the fully reduced states. Whereas the E(m) value of native azurin is pH dependent, the E(m) value of Cu(A) azurin is not, as expected for the Cu(A) center. Similarities and differences in the redox properties are discussed in terms of the known crystal structures of Cu(A) centers in cytochrome c oxidase and Cu(A) azurin.
Department of Biochemistry, The University of Mississippi, Medical Center, Jackson, MS 39216-4505, USA.
Dapeng
D
Sun
Xiaotang
X
Wang
Victor L
VL
Davidson
D001400:Azurin / Q000737:chemistry* / Q000235:genetics; D003300:Copper / Q000737:chemistry; D004563:Electrochemistry; D003576:Electron Transport Complex IV / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D015394:Molecular Structure; D010084:Oxidation-Reduction; D015202:Protein Engineering; D011550:Pseudomonas aeruginosa / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013053:Spectrophotometry
D011994:Recombinant Proteins; D001400:Azurin; D003300:Copper; D003576:Electron Transport Complex IV