PH-INDUCED CONFORMATIONAL TRANSITIONS OF THE PROPEPTIDE OF HUMAN CATHEPSIN-L - A ROLE FOR A MOLTEN GLOBULE STATE IN ZYMOGEN ACTIVATION

Synthesis of proteases as inactive zymogens is a very important mechan ism for the regulation of their activity. For lysosomal proteases prot eolytic cleavage of the propeptide is triggered by the acidic pH, By u sing fluorescence, circular dichroism, and NMR spectroscopy, we show t hat upon decreasing the pH from 6.5 to 3 the propeptide of cathepsin L loses most of the tertiary structure, but almost none of the secondar y structure is lost, Another partially structured intermediate, prone to aggregation, was identified between pH 6.5 and 4., The conformation , populated below pH 4, where the activation of cathepsin L occurs, is not completely unfolded and has the properties of molten globule, inc luding characteristic binding of the 1-anilinonaphthalene-8-sulfonic a cid. This pH unfolding of the propeptide parallels a decrease of its a ffinity for cathepsin L and suggests the mechanism for the acidic zymo gen activation. Addition of anionic polysaccharides that activate cath epsin L already at pH 5.5 unfolds the tertiary structure of the propep tide at this pH, Propeptide of human cathepsin L which is able to fold independently represents an evolutionary intermediate in the emergenc e of novel inhibitors originating from the enzyme proregions.