Oral Presentation 29th Annual Lorne Proteomics Symposium 2024

Using multi-omics to unravel amino acid uptake and utilisation of blood stage Plasmodium falciparum malaria (#28)

Ghizal Siddiqui 1 , Chris a MacRaild 1 , Carlo Giannangelo 1 , Katherine e Ellis 1 , amanda DePaoli 1 , peter scammells 2 , Sheena McGowan 3 , Darren Creek 1
  1. Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville Campus, Parkville, Victoria, Australia
  2. Medicinal Chemistry, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, Victoria, Australia
  3. Biomedicine Discovery Institute and Department of Microbiology, Monash University, Clayton, Victoria, Australia

Plasmodium falciparum parasite cause the majority of malaria mortality worldwide. P. falciparum parasites infect human red blood cells degrading large amount of host haemoglobin (Hb) and salvaging the resulting amino acids (aas) for its own use. The parasite also actively takes up aas from its external environment, including isoleucine, which is absent from Hb and the parasite is unable to synthesise de novo. This requirement for aa scavenging, especially Hb digestion, is a point of critical vulnerability for the parasite, and is targeted by many antimalarial drugs. We used a multi-omics workflow consisting of metabolomics, fluxomics (using stable-isotope-labelled aas) and peptidomics to identify the source of aas (Hb or external) and their utilisation into critical and unique metabolic processes of the parasite. Metabolomics analysis of isotope incorporation from labelled aas revealed activity in essential parasite metabolic reactions, and also in unexpected metabolic pathways. This included the identification of several labelled metabolites in the methionine salvage pathway that was not known to be present. Inhibition of Hb digestion with a range of protease inhibitors resulted in unique signatures of peptide intermediates, however, aa levels were unchanged. We demonstrated that related aas (charged, uncharged and hydrophobic) have similar uptake profiles within the parasite, suggesting the presence of previously uncharacterised aa transporters. Orthogonal label-free chemoproteomic methods based on thermal stability and limited proteolysis identified parasite and host proteins to mediate isoleucine transport. This hijacking of host cell function to access nutrients was further confirmed by identification of cross-linked peptides between the identified parasite and host proteins. Together, our multi-omics approach provides detailed analysis of aa uptake and utilisation by the parasite, opening new avenues for drug discovery.