Chemical modifcation and inactivation of rat liver arginase by N-bromosuccinimide: Reaction with his141

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Treatment of rat liver arginase with N-bromosuccinimide results in modification of six tryptophan residues per enzyme molecule and is accompanied by loss of catalytic activity (E. Ber and G. Muzynska (1979) Acta Biochim. Pol. 26, 103-114). In order to probe the chemistry of N-bromosuccinimide inactivation and the role of tryptophan residues in catalysis, the two tryptophan residues of rat liver arginase, Trp122 and Trp164, have been separately mutated to phenylalanine using site-directed mutagenesis of the protein expressed in Escherichia coli. Both single Trp→Phe mutant enzymes have kinetic parameters nearly identical to those for the wild-type enzyme. Treatment of native, wild-type, and each of the Trp→Phe mutant enzymes with N-bromosuccinimide results in loss of absorbance at 280 nm and is accompanied by a loss of catalytic activity. However, treatment of the wild-type enzyme with N-bromosuccinimide in the presence of the arginase inhibitors AG-hydroxy-L-arginine or the combination of L-ornithine and borate protects against inactivation, even though tryptophan residues are modified. Treatment of the H1O1N and H126N mutant arginases with N-bromosuccinimide also results in loss of catalytic activity and modification of tryptophan residues. In contrast, the H141N mutant arginase is not inactivated by N-bromosuccinimide, indicating that His141 is the critical target for the N-bromosuccinimide inactivation of the enzyme.

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Archives of Biochemistry and Biophysics





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This article was published in Archives of Biochemistry and Biophysics, Volume 327, Issue 1, Pages 107-112.

The published version is available at http://dx.doi.org/10.1006/abbi.1996.0098.

Copyright © 1996 Elsevier.

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