A large upregulation of the dipeptidyl aminopeptidase-1, DPAP-1, was detected in a quantitative proteomic study in which the parasite was subjected to treatment with antimalarial drugs. DPAPs are recognized to have a function in hemoglobin degradation in the erythrocytic stages of the malaria parasite, <i>Plasmodium falciparum</i>. DPAP-1 has been compared to the human ortholog cathepsin C in previous studies, concluding that both proteases belong to the same family of C-1 endo- and exopeptidases. An assay was developed using a fluorogenic substrate to test for the role of the upstream pathway initiator, cytochrome c, in the putative programmed cell death pathway. In order to test if DPAP-1 binds and is activated by cytochrome c, the expression and purification of DPAP-1 protease is needed. The inactive enzyme was expressed starting at residue 28, an aspartate that is part of the exclusion domain. The PCR product was cloned in the pMKL1 plasmid, which has a maltose binding protein and a TEV protease site. A his-tag was included to aid in purification and affinity studies with cytochrome c. The recombinant DPAP-1 protease will help expand the knowledge on its function and its role in<i> Plasmodium falciparum</i>’s apoptosis pathway.

Expression and purification of DPAP-1, a malaria protease, and its role in the putative apoptosis pathway of the parasite

249th National Meeting (2015)

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