Poster Presentation 23rd Annual Lorne Proteomics Symposium 2018

Peptidomics and metabolomics reveal the role of cysteine, aspartic and metallo proteases in the haemoglobin digestion pathway of the malaria parasite. (#90)

Ghizal Siddiqui 1 , Nyssa Drinkwater 2 , Amanda E De Paoli 1 , Sheena McGowan 2 , Darren J Creek 1
  1. Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville Campus, Parkville, Victoria, Australia
  2. Biomedicine Discovery Institute and Department of Microbiology, Monash University, Clayton, Victoria, Australia

The malaria parasite requires amino acids for protein synthesis and a major source of amino acids is via the digestion of host haemoglobin. Digestion of haemoglobin occurs within a specialised food vacuole via a semi-ordered proteolytic cascade that is mediated by different protease including aspartic, cysteine and metallo proteases. As haemoglobin digestion is essential for parasite survival within the red blood cell, understanding the exact interplay between the different classes of proteases within the food vacuole is crucial. In this study, we have developed combined metabolomics and modified proteomics workflows to identify the ‘signature peptide’ libraries that are generated as a result of specific inhibition of each protease class involved in haemoglobin digestion. The combined peptidomics analysis revealed that inhibition of specific proteases resulted in unique changes in the abundance of endogenous peptides. The results obtained were highly sensitive and could distinguish between different clans of the same class of protease. For example, specific inhibition of the metallo-aminopeptidases could distinguish unique peptide signatures between different types of aminopeptidases. Inhibition of the clan MA, M1 aminopeptidase resulted in accumulation of peptides containing basic residues, while specific inhibition of the clan MF, M17 aminopeptidase resulted in accumulation of hydrophobic peptides. This difference in selectivity is consistent with previously published in vitro studies of substrate specificity. In conclusion, this multi-platform approach provided an extensive coverage of endogenous peptides liberated during haemoglobin digestion within the parasite, and identified specific peptide signatures associated with inhibition of different classes of proteases involved in haemoglobin digestion. This will subsequently enhance our understanding of the interplay between the different proteases involved in haemoglobin digestion and reveal the mechanisms of action and resistance for drugs that target this pathway.