Protease pro region required for folding is a potent inhibitor of the mature enzyme

Abstract

alpha-Lytic protease, an extracellular bacterial serine protease, is synthesized with a large pro region that is required in vivo for the proper folding of the protease domain. To allow detailed mechanistic study, we have reconstituted pro region-dependent folding in vitro. The pro region promotes folding of the protease domain in the absence of other protein factors or exogenous energy sources. Surprisingly, we find that the pro region is a high affinity inhibitor of the mature protease. The pro region also inhibits the closely related Streptomyces griseus protease B, but not the more distantly related, yet structurally similar protease, elastase. Based on these data, we suggest a mechanism in which pro region binding reduces the free energy of a late folding transition state having native-like structure.

Intramolecular inhibition of human defensin HNP-1 by its propiece.

“…The common occurrence of relatively large proregions that are degraded during posttranslational processing argues that they fulfill important roles during protein biosynthesis and subcellular transport. Studies of proteolytic enzymes indicate that the proregion may catalyze correct protein folding (23,24) or act as a chaperone that prevents promiscuous protein-protein interactions. In some hormones the proregion contains a targeting signal for subcellular sorting (25) thus freeing the mature peptide from the constraints of this function.…”

Inhibition of cathepsin B by its propeptide: Use of overlapping peptides to identify a critical segment

“…The proregions of these proteinases are essential for such physiological functions as intracellular trafficking, correct folding of the enzyme during maturation, and inhibition of the mature enzyme [1][2][3][4][5]. The inhibition of proteinases by their respective proregions is a rather specific process [6,7], and this relationship can be used to develop a new generation of specific peptide inhibitors. The cysteine proteinases of the papain family are particularly appropriate targets for such a strategy, since no specific synthetic substrates and inhibitors are available to follow the activity of individual members.…”

Specific Inhibition and Stabilization of Aspergilloglutamic Peptidase by the Propeptide

“…These roles, however, have not yet been fully understood, and further studies on various proteins are necessary. As for the inhibitory properties of the propeptides of peptidases, there are numbers of reports describing their inhibition profiles and type of inhibition (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25). However, only a few attempts have been made so far to identify critical sequences and/or residues in the propeptide important for inhibition of the corresponding mature peptidase, and much of the inhibition mechanisms, the structural determinants in the propeptide, and the sites of binding involved in the inhibition remains to be established.…”