A glycoengineered antigen exploiting a conserved protein O-glycosylation pathway in the Burkholderia genus for detection of glanders infections.
A glycoengineered antigen exploiting a conserved protein O-glycosylation pathway in the Burkholderia genus for detection of glanders infections.
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DOI：
10.1080/21505594.2021.1876440
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发表时间：
2021-12
期刊：
影响因子：
5.2
通讯作者：
Valvano MA
中科院分区：
文献类型：
--
作者：
Wang G;Glaser L;Scott NE;Fathy Mohamed Y;Ingram R;Laroucau K;Valvano MA
我们最近描述了一种蛋白质O-糖基化途径保守在所有物种的伯克霍尔德氏菌属，导致在合成和掺入的三糖聚糖膜输出蛋白。在这里，我们利用这个系统来构建和评估一个诊断工具的鼻疽。鼻疽的病原体鼻疽伯克霍尔德菌是一种高度传染性和致命的人畜共患病原体，感染马，骡子，驴，偶尔也感染人类。高度敏感和特异性的诊断工具对于控制、消除和根除B至关重要。鼻疽感染我们构建了携带合成基因的质粒，该基因编码一种经修饰的、先前未注释的伯克霍尔德氏菌糖蛋白，该糖蛋白含有三个与霍乱毒素B亚基融合的糖基化序列。所得蛋白质在B中糖基化。通过SDS-PAGE和荧光凝集素印迹法测定，新洋葱K56-2亲本菌株中的糖基化水平高于亲本菌株，但糖基化缺陷突变体中的糖基化水平低于亲本菌株。这些糖蛋白之一被用作ELISA和蛋白质印迹法中的抗原，以筛选从腺感染和健康马收集的一组血清样品，这些血清样品先前通过补体结合试验和间接ELISA基于B的半纯化级分进行了研究。鼻疽我们表明，基于我们的糖蛋白抗原的ELISA和蛋白质印迹分析提供了100%的特异性，灵敏度大于88%。从感染B的患者收集的血清样品识别糖蛋白抗原。类鼻疽B. B.鼻疽B. mallei，B.多噬菌体和B. cenocepacia。我们的研究结果表明，蛋白质O-糖基化伯克霍尔德氏菌可以利用作为诊断伯克霍尔德氏菌相关感染的生物标志物。
We recently described a protein O-glycosylation pathway conserved in all species of the Burkholderia genus that results in the synthesis and incorporation of a trisaccharide glycan to membrane-exported proteins. Here, we exploited this system to construct and evaluate a diagnostic tool for glanders. Burkholderia mallei, the causative agent of glanders, is a highly infectious and fatal zoonotic pathogen that infects horses, mules, donkeys, and occasionally humans. A highly sensitive and specific diagnostic tool is crucial for the control, elimination, and eradication of B. mallei infections. We constructed plasmids carrying synthetic genes encoding a modified, previously unannotated Burkholderia glycoprotein containing three glycosylation sequons fused to the cholera toxin B-subunit. The resulting proteins were glycosylated in the B. cenocepacia K56-2 parental strain, but not in glycosylation-deficient mutants, as determined by SDS-PAGE and fluorescent lectin blots. One of these glycoproteins was used as an antigen in ELISA and western blots to screen a panel of serum samples collected from glanders-infected and healthy horses, which were previously investigated by complement fixation test and indirect ELISA based on a semi-purified fraction of B. mallei. We show that ELISA and western blot assays based on our glycoprotein antigen provide 100% specificity, with a sensitivity greater than 88%. The glycoprotein antigen was recognized by serum samples collected from patients infected with B. pseudomallei, B. mallei, B. multivorans, and B. cenocepacia. Our results indicate that protein O-glycosylation in Burkholderia can be exploited as a biomarker for diagnosis of Burkholderia-associated infections.
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影响因子：
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作者：
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通讯作者：
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作者：
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