Figure 5.

Autoradiogram of extract from radiolabeled [¹⁴C]Mu-F-hF-VSφ intracellular amastigotes. Binding of the radiolabeled inhibitor to amastigote cruzain was abolished by preincubation with unlabeled Mu-F-hF-VSφ followed by [¹⁴C]CPI. Epimastigote cruzain and recombinant cruzain controls are shown as standards. Note two species labeled in amastigotes that comigrate with either epimastigote cruzain (50 kD) which has both catalytic and COOH-terminal domains (13) or with recombinant cruzain (30 kD) that only has catalytic domain. Lane 1, T. cruzi intracellular amastigotes labeled with [¹⁴C]CPI; lane 2, T. cruzi intracellular amastigotes preincubated with unlabeled inhibitor; lane 3, sample buffer; lane 4, recombinant cruzain labeled with [¹⁴C]CPI; lane 5, epimastigotes radiolabeled with [¹⁴C]Mu-F-hF-VSφ. Note 57/51-kD doublet characteristic of native cruzain which retains COOH-terminal domain (13). Recombinant cruzain lacks the COOH-terminal domain (14).