T. gondii TIR1 strain temperature range thermal profiling experiment

ABSTRACT: Toxoplasma gondii is a single-celled eukaryotic parasite that chronically infects a quarter of the global population. In recent years, phenotypic screens have identified compounds that block parasite replication. Unraveling the complexity of pathogenesis and molecular mechanisms and pathways perturbed by such compounds requires target deconvolution. Thermal proteome profiling (TPP), also known as the cellular thermal shift assay (CETSA), recently emerged as a method to identify small molecule–target interactions in living cells and cell extracts in a variety of organisms, including unicellular eukaryotic pathogens. Ligand binding induces a thermal stability shift—stabilizing or destabilizing proteins that change conformationally in response to the ligand—that can be measured by mass spectrometry (MS). Cells are incubated with different concentrations of ligand and heated, causing thermal denaturation of proteins. The soluble protein is extracted and quantified with multiplexed, quantitative MS, giving rise to thousands of thermal denaturation profiles. Proteins engaging the ligand can be identified by their concentration-dependent thermal shift. The protocol provided here can be used to identify compound-target interactions and assess the impact of environmental or genetic perturbation on the thermal stability of the proteom in T. gondii and other eukaryotic pathogens. Here, we generate a reference dataset of the thermal profiles of extracellular T. gondii tachyzoites.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Toxoplasma Gondii Rh

SUBMITTER: Alice Herneisen  

LAB HEAD: Sebastian Lourido

PROVIDER: PXD024912 | Pride | 2023-03-11

REPOSITORIES: Pride