Poster Presentation The 16th Australian Peptide Conference 2025

Recombinant production of an antimalarial peptide in Nicotiana benthamiana (#239)

Linh T.T. Nguyen 1 , Maxim D. Harding 1 , Wing L. Healy 1 , Mengbai Zhang 1 , Nipuni Peththa Thanthrige 1 , Thao T. Ninh 2 , Mark A. Jackson 2 , Edward K. Gilding 1 , Nicole Lawrence 1 , David J. Craik 1
  1. Institute for Molecular Bioscience, Australian Research Council Centre of Excellence for Innovations in Peptide and Protein Science, The University of Queensland, Brisbane, Queensland, Australia
  2. Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, Brisbane, Queensland, Australia

Malaria is a life-threatening infectious disease with over half a million deaths recorded every year, a number that is expected to rise due to the spread of resistance to current antimalarial drugs. New drugs with different mechanisms of killing malaria parasites are urgently needed. To address this need, we are developing membrane-active peptides called PDIPs, that selectively enter infected red blood cells to disrupt parasite digestive vacuole membranes. PDIP analogues have been successfully generated using chemical synthesis; however, we aim to produce them in Nicotiana benthamiana as a more environmentally friendly and scalable approach. Positively charged and membrane-active peptides can be difficult to produce biosynthetically due to their inherent toxicity to host cells. We have overcome this challenge by expressing PDIP fused to calmodulin, a protein shown to bind other positively charged peptides, in N. benthamiana leaves. Further enhancements in PDIP accumulation were achieved by eliciting a heat-shock response in N. benthamiana plants, or by applying ascorbic acid to the leaves after infiltration with PDIP fusion constructs. We are currently optimising processing of plant material to liberate free PDIP, with the ultimate goal of employing a plant ligase enzyme that can be activated in planta or in vitro. This production platform represents a promising alternative for producing a wide range of bioactive positively charged peptides.