title: Global Profiling and Inhibition of Protein Lipidation in Vector and Host Stages of the Sleeping Sickness ParasiteTrypanosoma brucei.
abstract: The enzyme N-myristoyltransferase (NMT) catalyzes the essential fatty acylation of substrate proteins with myristic acid in eukaryotes and is a validated drug target in the parasiteTrypanosoma brucei, the causative agent of African trypanosomiasis (sleeping sickness). N-Myristoylation typically mediates membrane localization of proteins and is essential to the function of many. However, only a handful of proteins are experimentally validated as N-myristoylated inT. brucei. Here, we perform metabolic labeling with an alkyne-tagged myristic acid analogue, enabling the capture of lipidated proteins in insect and host life stages ofT. brucei. We further compare this with a longer chain palmitate analogue to explore the chain length-specific incorporation of fatty acids into proteins. Finally, we combine the alkynyl-myristate analogue with NMT inhibitors and quantitative chemical proteomics to globally define N-myristoylated proteins in the clinically relevant bloodstream form parasites. This analysis reveals five ARF family small GTPases, calpain-like proteins, phosphatases, and many uncharacterized proteins as substrates of NMT in the parasite, providing a global view of the scope of this important protein modification and further evidence for the crucial and pleiotropic role of NMT in the cell.
authors.affiliation: Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
authors.forename: Megan H
authors.initials: MH
authors.lastname: Wright
authors.affiliation: Centre for Immunology and Infection, Department of Biology, University of York , York YO10 5DD, United Kingdom.
authors.forename: Daniel
authors.initials: D
authors.lastname: Paape
authors.affiliation: Centre for Immunology and Infection, Department of Biology, University of York , York YO10 5DD, United Kingdom.
authors.forename: Helen P
authors.initials: HP
authors.lastname: Price
authors.affiliation: Centre for Immunology and Infection, Department of Biology, University of York , York YO10 5DD, United Kingdom.
authors.forename: Deborah F
authors.initials: DF
authors.lastname: Smith
authors.affiliation: Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
authors.forename: Edward W
authors.initials: EW
authors.lastname: Tate
keywords: N-myristoylation; chemical proteomics; click chemistry; human African trypanosomiasis; protein lipidation; target validation