Mechanism of activation of trypsinogen and chymotrypsinogen

The Prodomain of Ssy5 Protease Controls Receptor-Activated Proteolysis of Transcription Factor Stp1

“…An important characteristic of RAP is that Ssy5 autolysis, which occurs concomitantly with its biogenesis, is not sufficient to trigger Stp1 processing (3). In this respect, Ssy5 differs from classical chymotrypsin-like proteases that are most commonly activated by intermolecular proteolytic processing of an inactive zymogen precursor (32). The processing of the prodomain within the zymogen is frequently associated with a conformational change that suffices to fully activate the protease (18,41,45).…”

Autoactivation of Thrombin Precursors

“…Conformational selection in terms of the pre-existing E*-E equilibrium provides a more realistic paradigm for the function of the zymogen and protease and challenges the current dogma of the zymogen as an inactive precursor of the active protease (4,5). The molecular basis of autoactivation in terms of conformational selection also defines a strategy for the facile production of trypsin-like proteases of clinical and biotechnological relevance.…”

“…The zymogen is proteolytically cut at Arg-15 in nearly all members of the family to generate a new N terminus that ion-pairs with the highly conserved Asp-194 next to the catalytic Ser-195 and organizes both the oxyanion hole and primary specificity pocket for substrate binding and catalysis (3). Central to this paradigm is that the zymogen does not convert spontaneously to the mature enzyme, which in turn does not feed back to activate its zymogen form (4,5). A safety mechanism is thereby established to guarantee stability of the zymogen until a biological signal triggers activation via a distinct protease, a trypsin-like enzyme with primary specificity toward Arg side chains, which often acts in tandem with a cofactor in the context of a proteolytic cascade (6).…”

Kinetic Dissection of the Pre-existing Conformational Equilibrium in the Trypsin Fold

“…This scenario has physiological relevance in the context of autoactivation (8,9,28,86). The long accepted paradigm where the zymogen is viewed simply as the inactive form of the mature protease (30,31,87,88) needs revision (8). Zymogen and protease are two different incarnations of the underlying plasticity of the trypsin fold and are each capable of assuming closed (E*) and open (E) conformations of the active site.…”

“…In the case of the mature enzyme, the irreversible catalytic conversion of substrate into product that follows the binding step complicates resolution of the kinetic rates underscoring the mechanism of recognition (15). Furthermore, rapid kinetics of substrate recognition have historically focused on the steps that follow the initial binding step into the catalytic cycle (30,31). The E*-E equilibrium of the trypsin fold refocuses attention on the steps that precede the binding interaction and requires an experimental approach where catalysis is eliminated by suitable amino acid replacements that do not perturb the recognition event.…”