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PubMed · 1369354

Protein stabilization by engineered metal chelation.

J T Kellis·R J Todd·F H Arnold
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Abstract

A ligand can shift a protein's folding/unfolding equilibrium by binding with higher affinity to the native state. A metal-chelating site consisting of two histidines separated by three residues (His-X3-His) engineered into an alpha-helix provides a general and easily-implemented means for protein stabilization by this mechanism. We have tested this approach with the iso-1-cytochrome c of Saccharomyces cerevisiae substituted with histidine at positions 4 and 8 in its N-terminal alpha-helix. One mM Cu(II) complexed to iminodiacetate stabilizes the cytochrome c variant by ca. 1 kcal/mol, as determined by guanidinium chloride-induced unfolding. The protein's folding/unfolding equilibrium is shifted by a free energy equal to that calculated from the metal ion's preferential binding to the native protein. Given the ubiquity of surface alpha-helices and the additional possibility of inserting di-histidine chelating sites into turns and beta-structures, we conclude that this is a useful method for protein stabilization.

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Binding SitesChelating AgentsCytochrome c GroupCytochromes cDrug StabilityHistidineMetalsProtein ConformationProtein DenaturationProteinsSaccharomyces cerevisiaeSaccharomyces cerevisiae ProteinsJournal ArticleResearch Support, Non-U.S. Gov'tResearch Support, U.S. Gov't, Non-P.H.S.Research Support, U.S. Gov't, P.H.S.

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J T Kellis, R J Todd, F H Arnold. 1991. Protein stabilization by engineered metal chelation.. https://doi.org/10.1038/nbt1091-994

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