Proteomic analysis of the cyst stage of Entamoeba histolytica.
Proteomic analysis of the cyst stage of Entamoeba histolytica.
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DOI：
10.1371/journal.pntd.0001643
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发表时间：
2012
影响因子：
3.8
通讯作者：
Petri WA Jr
中科院分区：
文献类型：
--
作者：
Ali IK;Haque R;Siddique A;Kabir M;Sherman NE;Gray SA;Cangelosi GA;Petri WA Jr
B类生物恐怖分子，溶组织内阿米巴有两个生命周期：感染性包囊阶段和侵袭性滋养体阶段。由于我们不能在体外有效地诱导囊化，我们对囊状结构的了解仍然有限。这也阻碍了我们开发专门针对囊肿的诊断工具的能力。三个主要目的是(1)鉴定包囊样本中的溶组织埃希氏菌蛋白，(2)丰富我们对包囊阶段的认识，(3)鉴定候选蛋白以开发包囊特异性诊断工具。用Percoll(梯度)纯化从感染者的粪便中提纯包囊。用高灵敏度的LC-MS/MS质谱仪(Orbitrap)鉴定包囊蛋白。共鉴定出417个非冗余的溶组织埃希氏菌蛋白质，其中195个蛋白质从未在滋养体来源的蛋白质组或表达序列标签(EST)数据集中检测到，这与包囊特异性一致。囊壁特异性糖蛋白Jacob、Jessie和几丁质酶均呈阳性。针对雅各布产生的抗体在粪便样本中识别出了包囊，并有可能作为诊断试剂使用。还鉴定了几种蛋白激酶、小GTP酶信号分子、DNA修复蛋白、表观遗传调节蛋白和表面相关蛋白。我们确定的蛋白质很可能是排泄物中含量最丰富的蛋白质之一，因此有望成为诊断靶点。这里产生的蛋白质组数据是第一次自然产生的溶组织埃希氏菌包囊，它们提供了对感染性包囊形式的重要见解。此外，还鉴定了许多独特的候选蛋白，这将有助于开发新的诊断工具来鉴定溶组织埃希氏菌包囊。用串联质谱仪对5份包囊阳性粪便标本中的溶组肠杆菌包囊蛋白进行鉴定。我们报告了417个非冗余的溶组织埃希氏菌蛋白质的鉴定，其中包括195个蛋白质，这些蛋白质在现有滋养体来源的蛋白质组或EST数据集中没有被鉴定，这与包囊的特异性一致。由于这些包囊直接来自不完全纯化的患者样本，因此鉴定出数量有限的蛋白质(N = 417)，它们可能只代表部分蛋白质组。然而，这项研究成功地识别出在寄生虫的包囊阶段可能富含的蛋白质。这些蛋白中的几个可能在溶组织埃希氏菌的阶段转换或包囊功能中发挥作用。本研究中鉴定的蛋白质可能是诊断溶组织埃希氏菌包囊的有用标志物。总体而言，这项研究中产生的数据有望帮助了解寄生虫的包囊阶段，这对疾病传播和组织溶解埃希氏菌的发病机制至关重要。
The category B agent of bioterrorism, Entamoeba histolytica has a two-stage life cycle: an infective cyst stage, and an invasive trophozoite stage. Due to our inability to effectively induce encystation in vitro, our knowledge about the cyst form remains limited. This also hampers our ability to develop cyst-specific diagnostic tools. Three main aims were (i) to identify E. histolytica proteins in cyst samples, (ii) to enrich our knowledge about the cyst stage, and (iii) to identify candidate proteins to develop cyst-specific diagnostic tools. Cysts were purified from the stool of infected individuals using Percoll (gradient) purification. A highly sensitive LC-MS/MS mass spectrometer (Orbitrap) was used to identify cyst proteins. A total of 417 non-redundant E. histolytica proteins were identified including 195 proteins that were never detected in trophozoite-derived proteomes or expressed sequence tag (EST) datasets, consistent with cyst specificity. Cyst-wall specific glycoproteins Jacob, Jessie and chitinase were positively identified. Antibodies produced against Jacob identified cysts in fecal specimens and have potential utility as a diagnostic reagent. Several protein kinases, small GTPase signaling molecules, DNA repair proteins, epigenetic regulators, and surface associated proteins were also identified. Proteins we identified are likely to be among the most abundant in excreted cysts, and therefore show promise as diagnostic targets. The proteome data generated here are a first for naturally-occurring E. histolytica cysts, and they provide important insights into the infectious cyst form. Additionally, numerous unique candidate proteins were identified which will aid the development of new diagnostic tools for identification of E. histolytica cysts. We used tandem mass spectrometry to identify E. histolytica cyst proteins in 5 cyst positive stool samples. We report the identification of 417 non-redundant E. histolytica proteins including 195 proteins that were not identified in existing trophozoite derived proteome or EST datasets, consistent with cyst specificity. Because the cysts were derived directly from patient samples with incomplete purification, a limited number of proteins were identified (N = 417) that probably represent only a partial proteome. Nevertheless, the study succeeded in identifying proteins that are likely to be abundant in the cyst stage of the parasite. Several of these proteins may play roles in E. histolytica stage conversion or cyst function. Proteins identified in this study may be useful markers for diagnostic detection of E. histolytica cysts. Overall, the data generated in this study promises to aid the understanding of the cyst stage of the parasite which is vital for disease transmission and pathogenesis in E. histolytica.
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影响因子：
1.5
作者：
Das, Suchismita;Van Dellen, Katrina;Samuelson, John
通讯作者：
Samuelson, John
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2.1
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Bruchhaus, Iris
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7.7
作者：
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Mora-Galindo, J