AbstractLeishmania (Leishmania) amazonensis is an important agent of cutaneous leishmaniasis in Brazil. This parasite faces cell death in some situations during transmission to the vertebrate host, and this process seems to be dependent on the activity of metacaspase (MCA), an enzyme bearing trypsin-like activity present in protozoans, plants and fungi. In fact, the association between MCA expression and cell death induced by different stimuli has been demonstrated for several Leishmania species. Regulators and natural substrates of MCA are poorly known. To fulfill this gap, we have employed phage display over recombinant L. (L.) amazonensis MCA to identify peptides that could interact with the enzyme and modulate its activity. Four peptides were selected for their capacity to specifically bind to MCA and interfere with its activity. One of these peptides, similar to ecotin-like ISP3 of L. (L.) major, decreases trypsin-like activity of promastigotes under heat shock, and significantly decreases parasite heat shock-induced death. These findings indicate that peptide ligands identified by phage display affect trypsin-like activity and parasite death, and that an endogenous peptidase inhibitor is a possible natural regulator of the enzyme.