Functions of Propeptide Parts in Cysteine Proteases

Abstract

Regulation of proteolytic enzyme activity is an essential requirement for cells and tissues because proteolysis at the wrong time and location may be lethal. Two principal mechanisms to control the activity of proteases have been developed during evolution. The first is the co-evolution of endogenous inhibitors, typically occurring in cellular compartments separated from those containing active enzymes. The second is the fact that proteases are synthesized as inactive or less active precursor molecules. They are activated, in some cases, upon an appropriate signal like acidification, Ca(++) -binding or, in other cases, by limited intra- or intermolecular proteolysis cleaving off an inhibitory peptide. These regulatory proenzyme regions have attracted much attention during the last decade, since it became obvious that they harbour much more information than just triggering activation. In this review we summarize experimental data concerning three functions of propeptides of clan CA family C1 cysteine peptidases (papain family), namely the selectivity of their inhibitory potency, the participation in correct intracellular targeting and assistance in folding of the mature enzyme. Cysteine peptidases of the CA-C1 family include members from the plant kingdom like papain as well as from the animal kingdom like the lysosomal cathepsins L and B. As it will be shown, the functions are determined by certain structural motifs conserved over millions of years after the evolutionary trails have diverged. The function of propeptides of two other important classes of cysteine peptidases - the calpains, clan CA family C4, and the caspases, clan CD family C 14 - are not considered in this review.

Improving the secretory capacity of Chinese hamster ovary cells by ectopic expression of effector genes: Lessons learned and future directions

“…Some secreted proteins are expressed as inactive proprotein precursors containing one or more inhibitory propeptides that need to be proteolytically cleaved off by propeptidases before full activity is achieved (Wiederanders et al, 2003). Since these propeptides in general are essential for protein folding (Chen and Inouye, 2008), propeptide-containing r-proteins need to be expressed as proproteins to prevent misfolding and degradation.…”

The Major Secreted Cathepsin L1 Protease of the Liver Fluke, Fasciola hepatica

“…Prosegments function as regulators of activity by binding to the substrate cleft in a reverse substrate mode (9 -11), and they act as intermolecular chaperones that are crucial to protein folding (12)(13)(14) and intracellular trafficking (15)(16)(17). Cleavage between the prosegment region and the catalytic domain is necessary to generate the active mature enzyme and usually occurs in acidic environments, whether that be intracellularly (9, 18 -21) or extracellularly (10,(22)(23)(24)(25). Various mammalian cathepsins, including cathepsins L (9, 10, 18, 26 -29), S (10, 30 -32), K (10,33,34), and B (9, 24 -27), autoactivate in vitro under acidic and reducing conditions.…”

“…Cleavage between the prosegment region and the catalytic domain is necessary to generate the active mature enzyme and usually occurs in acidic environments, whether that be intracellularly (9, 18 -21) or extracellularly (10,(22)(23)(24)(25). Various mammalian cathepsins, including cathepsins L (9, 10, 18, 26 -29), S (10, 30 -32), K (10,33,34), and B (9, 24 -27), autoactivate in vitro under acidic and reducing conditions. The F. hepatica cathepsin L zymogen autoactivates within the slightly acidic pH of the parasite gut lumen and, similarly to the mammalian enzymes, autoactivates in vitro at low pH (2).…”

“…The initial "trigger" that initiates activation still remains elusive (15) but may be the disruption of a salt bridge involving Asp Ϫ36 (papain numbering) of the prosegment, contained within a conserved GXNXFXD motif, as the pH decreases (35). Subsequent removal of the prosegment involves possible intramolecular and/or intermolecular cleavages that take place within the central portion of the prosegment and then at the C-terminal region of the prosegment close to the N terminus of the mature enzyme (9,10,35).…”

“…The high B-factors observed in this portion of the proenzyme structure are believed to facilitate proteolytic cleavage at this site (9,10), and several studies have shown that this region is particularly susceptible to trans-cleavage with the same or different protease (18,25,28,32,33). As a consequence of this lack of structural stability, within the papain superfamily, the C-terminal part of prosegments show little or no conservation (9).…”