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ACS Infect Dis. 2016 Jun 10;2(6):427-441. Epub 2016 Apr 29.

Global Profiling and Inhibition of Protein Lipidation in Vector and Host Stages of the Sleeping Sickness Parasite Trypanosoma brucei.

Author information

1
Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
2
Centre for Immunology and Infection, Department of Biology, University of York , York YO10 5DD, United Kingdom.

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 parasite Trypanosoma 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 in T. 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 of T. 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.

KEYWORDS:

N-myristoylation; chemical proteomics; click chemistry; human African trypanosomiasis; protein lipidation; target validation

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