Blood flukes of the genus Schistosoma cause the disease schistosomiasis that infects over 200 million people worldwide. Treatment relies on just one drug, and new therapies are needed should drug resistance emerge. Schistosoma mansoni cathepsin B1 (SmCB1) is a gut-associated protease that digests host blood proteins as source of nutrients. It is under evaluation as a therapeutic target. Enzymatic activity of the SmCB1 zymogen is prevented by the pro-peptide that sterically blocks the active site until activation of the zymogen to the mature enzyme. We investigated the structure-inhibition relationships of how the SmCB1 pro-peptide interacts with the enzyme core using a SmCB1 zymogen model and pro-peptide-derived synthetic fragments. Two regions were identified within the pro-peptide that govern its inhibitory interaction with the enzyme core: an "active site region" and a unique "heparin-binding region" that requires heparin. The latter region is apparently only found in the pro-peptides of cathepsins B associated with the gut of trematode parasites. Finally, using the active site region as a template and a docking model of SmCB1, we designed a series of inhibitors mimicking the pro-peptide structure, the best of which yielded low micromolar inhibition constants. Overall, we identify a novel glycosaminoglycan-mediated mechanism of inhibition by the pro-peptide that potentially regulates zymogen activation and describe a promising design strategy to develop antischistosomal drugs.

Institute of Organic Chemistry and Biochemistry Academy of Sciences of the Czech Republic Flemingovo nam 2 Prague Czech Republic

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