Cystatin from Austrelaps superbus snake venom as a model for identifying potential inhibitors of Trypanosoma cruzi cruzain.

BACKGROUND

Chagas disease (CD), caused by , affects approximately seven million individuals worldwide, with the highest number of cases in Latin America. CD has two phases, of which the chronic phase is characterized by reduced efficacy in drug therapies. This and other factors make developing new strategies that aim to identify molecules capable of becoming alternatives to or complement current chemotherapy vitally important.

METHODS

Cruzain and AsCystatin were obtained recombinantly through expression in . Bioinformatic assays were conducted with both molecules, followed by enzyme inhibition assays. Subsequently, studies allowed for the design of peptides, which were then assessed for molecular interactions with cruzain. The designed peptides were synthesized, and their inhibitory potential on cruzain and their trypanocidal and cytotoxic effects were finally assessed.

RESULTS

AsCystatin, a potential inhibitor of cysteine proteases, was identified from previously published scientific literature. assays suggested that AsCystatin interacts with key regions of cruzain, and was subsequently produced through heterologous expression, obtaining a protein with a high degree of purity. Next, the inhibition of AsCystatin on the activity of cruzain was assessed, observing that approximately 20 µM of cystatin could inhibit 50% of the catalytic activity of the recombinant enzyme. Based on the analysis performed previously, original, and modified peptides were designed and tested, which allowed for identifying four peptides with inhibitory capacity on the enzymatic activity of cruzain. Finally, three of these peptides showed trypanocidal activity on epimastigote forms of in models.

CONCLUSION

It was possible to identify AsCystatin and four peptides derived from this protein with inhibitory activity on cruzain, highlighting the trypanocidal effect of these peptides observed in assays.