Rüter Christian, Silva Mariana Ruiz, Grabowski Benjamin, Lubos Marie-Luise, Scharnert Julia, Poceva Marija, von Tils Dominik, Flieger Antje, Heesemann Jürgen, Bliska James B, Schmidt M Alexander
Institute of Infectiology - Center for Molecular Biology of Inflammation (ZMBE), University of Münster, Germany.
Institute of Infectiology - Center for Molecular Biology of Inflammation (ZMBE), University of Münster, Germany.
Int J Med Microbiol. 2014 May;304(3-4):444-51. doi: 10.1016/j.ijmm.2014.02.003. Epub 2014 Feb 18.
The Yersinia outer protein M (YopM) is a type 3 secretion system (T3SS)-dependent effector protein of Yersinia enterocolitica, Yersinia pseudotuberculosis and Yersinia pestis. Although YopM is indispensable for full virulence, its molecular functions still remain largely elusive. Recently, we could identify the recombinant YopM (rYopM) protein derived from the Y. enterocolitica strain 8081 (JB580) as a cell-penetrating protein, which down-regulates the expression of various pro-inflammatory cytokines including TNFα. In this study, we have generated rabbit monoclonal anti-YopM antibodies (RabMabs). RabMabs were characterized by SDS-PAGE and Western blotting using various truncated versions of rYopM to identify epitope-containing domains. RabMabs recognizing either the N- or C-terminus of YopM were characterized further and validated using a collection of 61 pathogenic and non-pathogenic Yersinia strains as well as exemplary strains of major intestinal bacterial pathogens such as Salmonella enterica ssp. enterica, Shigella flexneri and intestinal pathogenic Escherichia coli. RabMab 41.3 directed at the N-terminus of YopM of Y. enterocolitica strain 8081 recognized all YopM-expressing pathogenic Yersinia strains analyzed in this study but failed to recognize non-pathogenic isolates. Thus, RabMab 41.3 might be applicable for the detection of pathogenic Yersinia strains.
耶尔森氏菌外膜蛋白M（YopM）是小肠结肠炎耶尔森氏菌、假结核耶尔森氏菌和鼠疫耶尔森氏菌中一种依赖于Ⅲ型分泌系统（T3SS）的效应蛋白。尽管YopM对于完全毒力是不可或缺的，但其分子功能仍很大程度上不清楚。最近，我们能够鉴定出来自小肠结肠炎耶尔森氏菌8081菌株（JB580）的重组YopM（rYopM）蛋白是一种细胞穿透蛋白，它能下调包括TNFα在内的多种促炎细胞因子的表达。在本研究中，我们制备了兔抗YopM单克隆抗体（RabMabs）。通过SDS-PAGE和使用rYopM的各种截短形式进行蛋白质印迹来表征RabMabs，以鉴定含表位的结构域。进一步表征并使用61种致病性和非致病性耶尔森氏菌菌株以及主要肠道细菌病原体如肠炎沙门氏菌亚种肠炎沙门氏菌、福氏志贺氏菌和肠道致病性大肠杆菌的典型菌株验证了识别YopM N端或C端的RabMabs。针对小肠结肠炎耶尔森氏菌8081菌株YopM N端的RabMab 41.3识别了本研究中分析的所有表达YopM的致病性耶尔森氏菌菌株，但未能识别非致病性分离株。因此，RabMab 41.3可能适用于致病性耶尔森氏菌菌株的检测。
