Excretion/secretion products from Schistosoma mansoni adults, eggs and schistosomula have unique peptidase specificity profiles

Abstract

Schistosomiasis is one of a number of chronic helminth diseases of poverty that severely impact personal and societal well-being and productivity. Peptidases (proteases) are vital to successful parasitism, and can modulate host physiology and immunology. Interference of peptidase action by specific drugs or vaccines can be therapeutically beneficial. To date, research on peptidases in the schistosome parasite has focused on either the functional characterization of individual peptidases or their annotation as part of global genome or transcriptome studies. We were interested in functionally characterizing the complexity of peptidase activity operating at the host-parasite interface, therefore the excretory-secretory products of key developmental stages of Schistosoma mansoni that parasitize the human were examined. Using class specific peptidase inhibitors in combination with a multiplex substrate profiling assay, a number of unique activities derived from endo- and exo-peptidases were revealed in the excretory-secretory products of schistosomula (larval migratory worms), adults and eggs. The data highlight the complexity of the functional degradome for each developmental stage of this parasite and facilitate further enquiry to establish peptidase identity, physiological and immunological function, and utility as drug or vaccine candidates.

SmSP2: A serine protease secreted by the blood fluke pathogen Schistosoma mansoni with anti-hemostatic properties

“…The extract was cleared by centrifugation (16,000 g at 4°C for 10 min), ultra-filtered using a 0.22 μm Ultrafree-MC device (Millipore) and stored at -80°C. Excretory/secretory products (ESP) of adult worms were collected as described previously [ 29 ]. Specifically, fifty pairs of adult worms were washed five times in Basch medium 169 containing 5% fetal bovine serum, 100 U/mL penicillin, 100 μg/mL streptomycin and 1% Fungizone (Gibco), incubated for 1h at 37°C under a 5% CO 2 atmosphere, washed five times and then incubated overnight at 37°C in 5% CO 2 in the above medium supplemented with 5% fetal calf serum but in the absence of Fungizone.…”

“…The Multiplex Substrate Profiling by Mass Spectrometry (MSP-MS) assay was performed as previously described [ 29 ]. SmSP2, human plasma kallikrein (Sigma-Aldrich) and bovine trypsin Sigma-Aldrich) (all 17 nM) were incubated in triplicate with a mixture of 228 synthetic tetradecapeptides (0.5 uM each) in 10 mM Tris-HCl, pH 8.0.…”

Cysteine proteases during larval migration and development of helminths in their final host

“…Recent findings using a functional degradomics strategy on the excretory–secretory products (ESP) of S . mansoni eggs identified a clan CA cysteine protease with activity at neutral pH [ 5 ]. The possible functions of these egg proteases include yolk degradation (as found for similar proteases in insect eggs and in filarial worms, as described above), egg hatching, and/or facilitating the passage of eggs through host tissues [ 6 ].…”

Global substrate specificity profiling of post‐translational modifying enzymes

“…The MSP‐MS assay has been used with protease inhibitors, gene deletions, and immunodepletion in combination with traditional proteomic methods to identify component proteases that are highly active in complex biological samples. This has enabled the “deconvolution” of proteolytic signatures from fungal pathogens, parasitic organisms, cancer cell lines, and patient samples, and allowed for the prioritization of proteases based on their functional contribution to the global substrate specificity profile. For example, the MSP‐MS assay was used to analyze the global activity signatures of the opportunistic fungal pathogen Candida albicans in the planktonic and biofilm states .…”