Abstract

The protein profile of dormant Mtb obtained after the gradual acidification of Mtb culture was studied to find antigenic proteins for humans that are expressed by M. tuberculosis (Mtb) cells in vitro under conditions close to the situation of persistence in vivo. According to 2D electrophoresis, a significant diversity of proteins in dormant cells was found. However, the representation of individual proteins in dormant versus active cells differed substantially. Immunoblotting in different protein fractions of dormant cells revealed ten proteins that are able to bind antibodies in pooled sera of TB patients. Two proteins (Rv2018 and Rv0341) are new immunogenics that were not previously found in other studies. Four proteins (Rv0341, Rv2018, Rv1509, Rv2986) with the maximal structural specificity for Mtb due to their unique extended domains were selected for further analysis. These proteins were expressed in E. coli cells and studied via enzyme-linked immunosorbent assay (ELISA) for the immunogenicity of individual sera of TB patients and healthy donors. All proteins were found to have the ability to react with individual sera of TB patients. In TB patients, 5–45% (depending on the particulate protein) have a titer that is higher than the average titers of healthy donors +SD; the most immunogenic was protein Rv2986. Thus, the application of phenotypically changed (dormant) Mtb cells makes it possible to identify a specific repertoire of immunodominant proteins that could be used in the construction of polypeptides that are useful for the serodiagnosis of active/latent TB.

中文翻译：

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休眠的结核分枝杆菌细胞的免疫反应蛋白

摘要

研究了在Mtb培养物逐渐酸化后获得的休眠Mtb的蛋白质谱，以寻找结核分枝杆菌（Mtb）表达的人类抗原蛋白。）细胞在接近体内持续存在情况的条件下进行。根据二维电泳，在休眠细胞中发现了显着的蛋白质多样性。但是，在休眠细胞与活动细胞中单个蛋白质的表示形式却大不相同。在休眠细胞的不同蛋白质部分中进行的免疫印迹分析显示，十种蛋白质能够与结核病患者合并血清中的抗体结合。两种蛋白质（Rv2018和Rv0341）是新的免疫原性，以前在其他研究中未发现。选择了由于其独特的扩展域而对Mtb具有最大结构特异性的四种蛋白质（Rv0341，Rv2018，Rv1509，Rv2986）进行进一步分析。这些蛋白质在大肠杆菌中表达并通过酶联免疫吸附测定（ELISA）研究了TB患者和健康供体的单个血清的免疫原性。发现所有蛋白质都具有与结核病患者血清反应的能力。在结核病患者中，有5–45％（取决于颗粒蛋白）的滴度高于健康供体+ SD的平均滴度。最具免疫原性的是蛋白质Rv2986。因此，表型改变（休眠）的Mtb细胞的应用使得有可能鉴定免疫显性蛋白质的特定库，该蛋白质可用于构建可用于活动/潜在TB的血清学诊断的多肽。