Evidence for an Active-Center Histidine in Trypsin through Use of a Specific Reagent, 1-Chloro-3-tosylamido-7-amino-2-heptanone, the Chloromethyl Ketone Derived from Nα-Tosyl-L-lysine*
摘要：
The chloromethyl ketone derived from Nα-tosyl-L-lysine, TLCK (1-chloro-3-tosylamido-7-amino-2-heptanone), has been synthesized as an active-center labeling reagent for trypsin since it combines specificity features and a reactive grouping. It inactivated trypsin with a second-order rate constant of 5.6 1. mole-1 sec-1, at pH 6, but did not do so in the presence of a competitive inhibitor of trypsin, benzamidine, or in 8 M urea. These observations indicate that TLCK reacted at the active center of trypsin. With radioactive inhibitor the reaction was shown to be equimolar, and analysis revealed loss of one histidine residue. Chymotrypsin was unaffected.
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DOI：
10.1021/bi00886a039
被引量：
年份：
1965
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