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

TY - JOUR

T1 - Global Profiling and Inhibition of Protein Lipidation in Vector and Host Stages of the Sleeping Sickness Parasite Trypanosoma brucei

AU - Wright, Megan H

AU - Paape, Daniel

AU - Price, Helen P

AU - Smith, Deborah F

AU - Tate, Edward W

N1 - © 2016, American Chemical Society.

PY - 2016/6/10

Y1 - 2016/6/10

N2 - 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.

AB - 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.

KW - Journal Article

U2 - 10.1021/acsinfecdis.6b00034

DO - 10.1021/acsinfecdis.6b00034

M3 - Article

C2 - 27331140

SN - 1471-2334

VL - 2

SP - 427

EP - 441

JO - BMC Infectious Diseases

JF - BMC Infectious Diseases

IS - 6

ER -