Oxidation of buried cysteines is slow and an insignificant factor in the structural destabilization of staphylococcal nuclease caused by H2O2 exposure

by Kim, Yun Ho; Stites, Wesley Eugene

The oxidation of buried cysteine or methionine residues can destroy the enzyme activity of a protein by disrupting structure. Engineering in such an oxidatively triggered switch for enzyme activity would only be useful if the effects of substitution are relatively minor, while the effects of the oxidized side chain upon structure are significant and the oxidation relatively easy. To assess the feasibility of this strategy for controlling enzyme activity, the effects of such substitutions and their oxidation were studied in a well characterized model protein, staphylococcal nuclease. Stability and enzyme activity of the oxidized proteins was assessed and compared to the stability and enzyme activity of the unoxidized proteins. Cysteines were found to be generally well tolerated in buried positions but these mutants were not more destabilized than wild-type when oxidized. This shows that buried cysteines are difficult enough to oxidize that this is not likely to be a useful protein engineering strategy or a commonly used regulatory modification. Similar effects were observed for methionine.

Journal
Amino Acids
Volume
27
Issue
2
Year
2004
Start Page
175
ISBN/ISSN
1438-2199; 0939-4451
DOI
10.1007/s00726-004-0110-8