Mutations Arg 117 3 His and Asn 21 3 Ile in human trypsinogen-I have been recently associated with hereditary pancreatitis (HP). The Arg 117 3 His substitution is believed to cause pancreatitis by stabilizing trypsin against autolytic degradation, while the mechanism of action of Asn 21 3 Ile has been unknown. In an effort to understand the effect(s) of this mutation, Thr 21 in the highly homologous rat trypsinogen-II was replaced with Asn or Ile, and the recombinant zymogens and their active

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... ypsin forms were studied. Kinetic parameters of all three trypsins were comparable, and the active enzymes suffered autolysis at similar rates, indicating that neither catalytic properties nor proteolytic stability of trypsin are influenced by mutations at position 21. When incubated at pH 8.0, 37°C, pure zymogens underwent autoactivation with concomitant trypsinolytic degradation in a Ca 2؉ -dependent fashion. Thus, in the presence of 5 mM Ca 2؉ , autoactivation and digestion of the zymogens after Arg 117 and Lys 188 were observed, while in the presence of 1 mM EDTA autoactivation and cleavage at Lys 188 were reduced, and zymogenolysis at the Arg 117 site was enhanced. Overall rates of zymogen degradation in [Asn 21 ]-and [Ile 21 ]trypsinogens were higher in Ca 2؉ than in EDTA, while [Thr 21 ]trypsinogen demonstrated inverse characteristics. Remarkably, both in the presence and absence of Ca 2؉ , [Ile 21 ]trypsinogen exhibited significantly higher stability against autoactivation and proteolysis than zymogens with Asn 21 or Thr 21 . The observations suggest that autocatalytic trypsinogen degradation may be an important defense mechanism against excessive trypsin generation in the pancreas, and trypsinogen stabilization by the Asn 21 3 Ile mutation plays a role in the pathogenesis of HP.