Photogeneration and Quenching of Tryptophan Radical in Azurin

• Published in: The journal of physical chemistry. B 2015-07, Vol.119 (29), p.9438-9449
• Main Authors: Larson, Bethany C, Pomponio, Jennifer R, Shafaat, Hannah S, Kim, Rachel H, Leigh, Brian S, Tauber, Michael J, Kim, Judy E
• Format: Article
• Language: English
• Subjects: Azurin - chemistry; Azurin - genetics; cobalt; Cobalt - chemistry; copper; electron transfer; Electrons; mutants; Mutation; Photolysis; physical chemistry; proteins; Pseudomonas aeruginosa; Spectrum Analysis; tryptophan; Tryptophan - chemistry; tyrosine; Tyrosine - chemistry; zinc
• ISSN: 1520-6106; 1520-5207; 1520-5207
• DOI: 10.1021/jp511523z

Tryptophan and tyrosine can form radical intermediates that enable long-range, multistep electron transfer (ET) reactions in proteins. This report describes the mechanisms of formation and quenching of a neutral tryptophan radical in azurin, a blue-copper protein that contains native tyrosine (Y108 and Y72) and tryptophan (W48) residues. A long-lived neutral tryptophan radical W48• is formed upon UV-photoexcitation of a zinc­(II)-substituted azurin mutant in the presence of an external electron acceptor. The quantum yield of W48• formation (Φ) depends upon the tyrosine residues in the protein. A tyrosine-deficient mutant, ZnIIAz48W, exhibited a value of Φ = 0.080 with a Co­(III) electron acceptor. A nearly identical quantum yield was observed when the electron acceptor was the analogous tyrosine-free, copper­(II) mutant; this result for the ZnIIAz48W:CuIIAz48W mixture suggests there is an interprotein ET path. A single tyrosine residue at one of the native positions reduced the quantum yield to 0.062 (Y108) or 0.067 (Y72). Wild-type azurin with two tyrosine residues exhibited a quantum yield of Φ = 0.045. These data indicate that tyrosine is able to quench the tryptophan radical in azurin.