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Cu(II)‐Binding properties of a cytochrome c with a synthetic metal‐binding site: His‐X 3 ‐His in an α‐helix
Author(s) -
Todd Robert J.,
Van Dam Mariana E.,
Casimiro Danilo,
Haymore Barry L.,
Arnold Frances H.
Publication year - 1991
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.340100209
Subject(s) - metal , chemistry , cytochrome c , helix (gastropod) , binding site , stereochemistry , crystallography , cytochrome , chelation , inorganic chemistry , organic chemistry , enzyme , biochemistry , biology , ecology , snail , mitochondrion
A metal‐binding site consisting of two histidines positioned His‐X 3 ‐His in an α‐helix has been engineered into the surface of Saccharomyces cerevisiae iso‐1‐cytochrome c. The synthetic metal‐binding cytochrome c retains its biological activity in vivo. Its ability to bind chelated Cu(II) has been characterized by partitioning in aqueous two‐phase polymer systems containing a polymer‐metal complex, Cu(II)IDA‐PEG, and by metal‐affinity chromatography. The stability constant for the complex formed between Cu(II)IDA‐PEG and the cytochrome c His‐X 3 ‐His site is 5.3 × 10 4 M −1 , which corresponds to a chelate effect that contributes 1.5 kcal mol −1 to the binding energy. Incorporation of the His‐X 3 ‐His site yields a synthetic metal‐binding protein whose metal affinity is sensitive to environmental conditions that alter helix structure or flexibility.