A novel nanobody against urease activity of Helicobacter pylori
A novel nanobody against urease activity of Helicobacter pylori
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DOI：
10.1016/j.ijid.2013.02.015
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发表时间：
2013-09-01
影响因子：
8.4
通讯作者：
Zare, Hamed
中科院分区：
文献类型：
--
作者：
Ardekani, Leila Safaee;Gargari, Seyed Latif Mousavi;Zare, Hamed
背景：幽门螺杆菌感染与胃炎有关，在某些情况下与胃和十二指肠溃疡，甚至腺癌有关。抗生素治疗有很大的局限性，如成本高和耐药性菌株的出现，产生了对新治疗方法的需求。抗H. pylori是一种新的有效的治疗策略。本研究成功构建了抗重组UreC的单可变区重链抗体。方法：以骆驼重链抗体为抗原，构建VHH噬菌粒文库。将纳米抗体展示在M13噬菌体上。针对UreC重组蛋白进行文库选择。在五轮淘选中筛选针对UreC的重链抗体的特异性单可变区。选择在噬菌体ELISA中具有最高得分的纳米抗体用于可溶性表达。用镍-次氮基三乙酸（Ni-NTA）柱纯化纳米抗体，并用十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）和Western印迹法进行确认。结果：成功分离纯化了一种特异性的UreC纳米抗体，该抗体对UreC重组蛋白具有高亲和力，可抑制尿素酶活性。结论：所制备的UreC纳米抗体能够特异性地检测并结合UreC，抑制尿素酶活性。这种纳米抗体可能是一种新型的治疗措施，针对H。幽门感染(C)2013年国际传染病学会。由爱思唯尔有限公司出版。保留所有权利。
Background: Helicobacter pylori infection is associated with gastritis and in some cases with gastric and duodenal ulcers, and even adenocarcinoma. Antibiotic therapy has significant limitations, such as the high cost and the emergence of antibiotic-resistant strains, generating the need for new treatments. The administration of antibody against H. pylori is a new effective therapeutic strategy. In this study, we successfully developed a single-variable domain of heavy chain antibody against recombinant UreC.Methods: A VHH phagemid library was constructed from immune camel heavy chain antibodies. The nanobodies were displayed on M13 phage. Library selection was performed against UreC recombinant protein. A specific single-variable domain of heavy chain antibody against UreC was screened in five rounds of panning. The nanobody with the highest score in the phage ELISA was selected for soluble expression. The nanobody was purified with a nickel-nitrilotriacetic acid (Ni-NTA) column and confirmed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Affinity, specificity, and urease inhibitory properties of the nanobody were assayed.Results: Here we showed the isolation and purification of a specific nanobody with high affinity against UreC recombinant protein that can inhibit urease activity.Conclusions: The isolated UreC nanobody can specifically detect and bind to UreC and inhibit urease activity. This nanobody could be a novel class of treatment measure against H. pylori infection. (C) 2013 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.