Fig. 2: Peptidases identified by mass spectrometry are released from intact trypanosomes.

a Release of identified peptidases by bloodstream form parasites. In each case, the identified peptidase was epitope tagged using constructs targeting the endogenous gene locus to ensure close to physiological expression. Resultant cell lines were assayed in serum-free Creek’s medium for 2 h and then the cell pellet (P) and supernatant (S) isolated by centrifugation. Isolated proteins were monitored for the distribution of each peptidase using the BB2 antibody detecting the Ty1 epitope tag, with the abundant cytosolic protein EF1-alpha used to monitor non-specific escape of cytosolic proteins. Molecular weight markers in kDa. b Inducible ectopic expression of each peptidase identified as released by parasites. In each case, N-terminally epitope-tagged genes were cloned into a doxycycline-inducible expression vector and then the expression and extracellular release of the expressed proteins were monitored using the epitope tag-specific antibody BB2, with the cytosolic protein EF1-alpha used as a control. Released and cell-associated proteins were isolated after incubation of parasites in serum-free Creek’s medium for 2 h and centrifugation. Molecular weight markers in kDa. Source data are provided as a source data file.