Woodward's reagent K reacts with histidine and cysteine residues in Escherichia coli and Saccharomyces cerevisiae phosphoenolpyruvate carboxykinases.
The reaction of Woordward's reagent K (WRK) with model amino acids and proteins has been analyzed. Our results indicate that WRK forms 340-nm-absorbing adducts with sulfhydryl- and imidazol-containing compounds, but not with carboxylic acid derivatives, in agreement with Liamas et al. [(1986), J. Am. Chem. Soc. 108, 5543-5548], but not with Sinha and Brewer [(1985), Anal. Biochem. 151, 327-333]. The chemical modification of Escherichia coli and Saccharomyces cerevisiae phosphoenolpyruvate carboxykinases with WRK leads to an increase in the absorption at 340 nm, and we have demonstrated its reaction with His and Cys residues in these proteins. These results caution against claims of glutamic or aspartic acid modification by WRK based on the absorption at 340 nm of protein- WRK adducts.
Departamento de Ciencias Químicas, Facultad de Química y Biología, Universidad de Santiago de Chile, Chile.
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Bustos
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Gajardo
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Gómez
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Goldie
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D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry*; D004926:Escherichia coli / Q000201:enzymology; D006639:Histidine / Q000737:chemistry*; D007202:Indicators and Reagents / Q000737:chemistry*; D007555:Isoxazoles / Q000737:chemistry*; D007700:Kinetics; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000737:chemistry*; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013056:Spectrophotometry, Ultraviolet
D000596:Amino Acids; D007202:Indicators and Reagents; D007555:Isoxazoles; C009686:N-ethyl-5-phenylisoxazolium-3'-sulfonate; D006639:Histidine; D010729:Phosphoenolpyruvate Carboxykinase (GTP); D003545:Cysteine