A Target Repurposing Approach Identifies N-myristoyltransferase as a New Candidate Drug Target in Filarial Nematodes
A Target Repurposing Approach Identifies N-myristoyltransferase as a New Candidate Drug Target in Filarial Nematodes
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DOI：
10.1371/journal.pntd.0003145
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发表时间：
2014-09-01
影响因子：
3.8
通讯作者：
Carlow, Clotilde K. S.
中科院分区：
文献类型：
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作者：
Galvin, Brendan D.;Li, Zhiru;Carlow, Clotilde K. S.
肉豆蔻酰化是一种脂质修饰，涉及在蛋白质亚群的n端甘氨酸上添加一种14碳不饱和脂肪酸，肉豆蔻酸，这种修饰促进它们与细胞膜结合以实现各种生物功能。该过程由肉豆蔻酰基辅酶a催化：蛋白质n -肉豆蔻酰基转移酶（NMT），该酶已被证实为人类癌症的药物靶点，以及由真菌、病毒和原生动物寄生虫引起的传染病。我们纯化了秀丽隐杆线虫和马来Brugia malayi NMTs作为活性重组蛋白，并对它们的必需脂肪酸供体、肉豆蔻酰基辅酶a和肽底物进行了动力学分析。生化和结构分析均表明，线虫酶是典型的NMT酶，与原生动物NMT酶具有高度的保守性。针对原生动物NMT的抑制化合物抑制线虫NMT的IC50值为2.5 ~ 10 nM，分别对12.5 μ M的马来细丝虫和50 μ M的成虫以及25 μ M的秀丽隐杆线虫有抑制作用。秀丽隐杆线虫的RNA干扰和基因缺失进一步表明，NMT对线虫的生存至关重要。观察到的影响可能是由于几个下游靶蛋白的功能被破坏。利用生物信息学分析预测了马来芽孢杆菌NMT的潜在底物。我们的遗传和化学研究强调了肉豆蔻酰化在线虫功能蛋白合成中的重要性，并首次表明NMT是寄生线虫生存所必需的。这些结果表明，靶向NMT可能是开发针对线虫病（包括丝虫病）的化疗药物的有效途径。
Myristoylation is a lipid modification involving the addition of a 14-carbon unsaturated fatty acid, myristic acid, to the N-terminal glycine of a subset of proteins, a modification that promotes their binding to cell membranes for varied biological functions. The process is catalyzed by myristoyl-CoA: protein N-myristoyltransferase (NMT), an enzyme which has been validated as a drug target in human cancers, and for infectious diseases caused by fungi, viruses and protozoan parasites. We purified Caenorhabditis elegans and Brugia malayi NMTs as active recombinant proteins and carried out kinetic analyses with their essential fatty acid donor, myristoyl-CoA and peptide substrates. Biochemical and structural analyses both revealed that the nematode enzymes are canonical NMTs, sharing a high degree of conservation with protozoan NMT enzymes. Inhibitory compounds that target NMT in protozoan species inhibited the nematode NMTs with IC50 values of 2.5-10 nM, and were active against B. malayi microfilariae and adult worms at 12.5 mu M and 50 mu M respectively, and C. elegans (25 mu M) in culture. RNA interference and gene deletion in C. elegans further showed that NMT is essential for nematode viability. The effects observed are likely due to disruption of the function of several downstream target proteins. Potential substrates of NMT in B. malayi are predicted using bioinformatic analysis. Our genetic and chemical studies highlight the importance of myristoylation in the synthesis of functional proteins in nematodes and have shown for the first time that NMT is required for viability in parasitic nematodes. These results suggest that targeting NMT could be a valid approach for the development of chemotherapeutic agents against nematode diseases including filariasis.