Giardia fatty acyl-CoA synthetases as potential drug targets.
Giardia fatty acyl-CoA synthetases as potential drug targets.
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DOI：
10.3389/fmicb.2015.00753
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发表时间：
2015
影响因子：
5.2
通讯作者：
Zhu G
中科院分区：
文献类型：
--
作者：
Guo F;Ortega-Pierres G;Argüello-García R;Zhang H;Zhu G
由肠贾第鞭毛虫（同义词 G.兰伯利亚，G.十二指肠）引起的贾第鞭毛虫病是全世界腹泻寄生虫病的主要原因之一。尽管治疗贾第鞭毛虫病的药物有限，但人们仍担心某些患者的毒性和新出现的耐药性。通过数据挖掘基因组序列，我们观察到肠杆菌无法从头合成脂肪酸（FA）。然而，这种寄生虫有五种长链脂肪酰辅酶A合成酶（GiACS1至GiACS5）来激活从宿主中清除的FA。 ACS是一种必需酶，因为FA需要被激活形成酰基辅酶A硫酯才能进入后续代谢。在本研究中，我们进行了实验来探索某些 GiACS 酶是否可以作为贾第鞭毛虫的药物靶点。基于高通量数据集和蛋白质模型分析，我们最初研究了 GiACS1 和 GiACS2，因为根据之前报道的转录组数据，发现编码这两种酶的基因在不同的寄生虫生命周期阶段以及与宿主细胞相互作用时表达更加一致。这两种蛋白被克隆并表达为重组蛋白。生化分析表明，两者都对棕榈酸 (C16:0) 和肉豆蔻酸 (C14:0) 具有明显的底物偏好，并且对棕榈酸或 ATP 具有变构或 Michaelis-Menten 动力学。 ACS 抑制剂 Triacsin C 抑制两种酶的活性（GiACS1 的 IC50 = 1.56 μM，Ki = 0.18 μM，GiACS2 的 IC50 = 2.28 μM，Ki = 0.23 μM）和体外 G. Enteris 的生长（IC50 = 0.8 μM）。正如贾第虫进化特征所预期的那样，与人类对应物相比，两种贾第虫在整体折叠结构上都表现出差异。这些观察结果支持这样的观点，即一些 GiACS 酶可能被探索作为这种寄生虫的药物靶点。
Giardiasis caused by Giardia intestinalis (syn. G. lamblia, G. duodenalis) is one of the leading causes of diarrheal parasitic diseases worldwide. Although limited drugs to treat giardiasis are available, there are concerns regarding toxicity in some patients and the emerging drug resistance. By data-mining genome sequences, we observed that G. intestinalis is incapable of synthesizing fatty acids (FA) de novo. However, this parasite has five long-chain fatty acyl-CoA synthetases (GiACS1 to GiACS5) to activate FA scavenged from the host. ACS is an essential enzyme because FA need to be activated to form acyl-CoA thioesters before they can enter subsequent metabolism. In the present study, we performed experiments to explore whether some GiACS enzymes could serve as drug targets in Giardia. Based on the high-throughput datasets and protein modeling analyses, we initially studied the GiACS1 and GiACS2, because genes encoding these two enzymes were found to be more consistently expressed in varied parasite life cycle stages and when interacting with host cells based on previously reported transcriptome data. These two proteins were cloned and expressed as recombinant proteins. Biochemical analysis revealed that both had apparent substrate preference toward palmitic acid (C16:0) and myristic acid (C14:0), and allosteric or Michaelis–Menten kinetics on palmitic acid or ATP. The ACS inhibitor triacsin C inhibited the activity of both enzymes (IC50 = 1.56 μM, Ki = 0.18 μM for GiACS1, and IC50 = 2.28 μM, Ki = 0.23 μM for GiACS2, respectively) and the growth of G. intestinalis in vitro (IC50 = 0.8 μM). As expected from giardial evolutionary characteristics, both GiACSs displayed differences in overall folding structure as compared with their human counterparts. These observations support the notion that some of the GiACS enzymes may be explored as drug targets in this parasite.