Global profiling and inhibition of protein lipidation in vector and host stages of the sleeping sickness parasite Trypanosoma brucei
Tate, EW; Wright, MH; Paape, D; Price, H; Smith, DF
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.
|Acceptance Date||Apr 29, 2016|
|Publication Date||Apr 29, 2016|
|Journal||ACS Infectious Diseases|
|Publisher||American Chemical Society|
|Keywords||human African trypanosomiasis, N-myristoylation, chemical proteomics, click chemistry, protein lipidation, target validation|
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