Chemical pulldown combined with mass spectrometry to identify the molecular targets of antimalarials in cell-free lysates

Abstract

Summary: Here, we provide a protocol using chemical pulldown combined with mass spectrometry (LC-MS/MS) to identify drug targets in Plasmodium falciparum. This approach works upon the principle that a resin-bound inhibitor selectively binds its molecular target(s) in cell-free lysates. We describe the preparation of drug beads and P. falciparum lysate, followed by chemical pulldown, sample fractionation, and LC-MS/MS analysis. We then detail how to identify specifically bound proteins by comparing protein enrichment in DMSO-treated relative to drug-treated lysates via quantitative proteomics.For complete details on the use and execution of this protocol, please refer to Milne et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell Biology
• Cell culture
• Microbiology
• Molecular/Chemical Probes
• Protein Biochemistry
• Proteomics