Journal of the American Chemical Society 2012-12-05

Azurin as a protein scaffold for a low-coordinate nonheme iron site with a small-molecule binding pocket.

Matthew P McLaughlin, Marius Retegan, Eckhard Bill, Thomas M Payne, Hannah S Shafaat, Salvador Peña, Jawahar Sudhamsu, Amy A Ensign, Brian R Crane, Frank Neese, Patrick L Holland

Index: J. Am. Chem. Soc. 134(48) , 19746-57, (2012)

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Abstract

The apoprotein of Pseudomonas aeruginosa azurin binds iron(II) to give a 1:1 complex, which has been characterized by electronic absorption, Mössbauer, and NMR spectroscopies, as well as X-ray crystallography and quantum-chemical computations. Despite potential competition by water and other coordinating residues, iron(II) binds tightly to the low-coordinate site. The iron(II) complex does not react with chemical redox agents to undergo oxidation or reduction. Spectroscopically calibrated quantum-chemical computations show that the complex has high-spin iron(II) in a pseudotetrahedral coordination environment, which features interactions with side chains of two histidines and a cysteine as well as the C═O of Gly45. In the (5)A(1) ground state, the d(z(2)) orbital is doubly occupied. Mutation of Met121 to Ala leaves the metal site in a similar environment but creates a pocket for reversible binding of small anions to the iron(II) center. Specifically, azide forms a high-spin iron(II) complex and cyanide forms a low-spin iron(II) complex.