Fercher Christian, Probst Ines, Kohler Verena, Goessweiner-Mohr Nikolaus, Arends Karsten, Grohmann Elisabeth, Zangger Klaus, Meyer N Helge, Keller Walter
Institute of Molecular Biosciences, NAWI Graz, University of Graz, Austria.
Division of Infectious Diseases, University Medical Center Freiburg, Germany.
Sci Rep. 2016 Apr 22;6:24643. doi: 10.1038/srep24643.
Untreatable bacterial infections caused by a perpetual increase of antibiotic resistant strains represent a serious threat to human healthcare in the 21(st) century. Conjugative DNA transfer is the most important mechanism for antibiotic resistance and virulence gene dissemination among bacteria and is mediated by a protein complex, known as type IV secretion system (T4SS). The core of the T4SS is a multiprotein complex that spans the bacterial envelope as a channel for macromolecular secretion. We report the NMR structure and functional characterization of the transfer protein TraH encoded by the conjugative Gram-positive broad-host range plasmid pIP501. The structure exhibits a striking similarity to VirB8 proteins of Gram-negative secretion systems where they play an essential role in the scaffold of the secretion machinery. Considering TraM as the first VirB8-like protein discovered in pIP501, TraH represents the second protein affiliated with this family in the respective transfer operon. A markerless traH deletion in pIP501 resulted in a total loss of transfer in Enterococcus faecalis as compared with the pIP501 wild type (wt) plasmid, demonstrating that TraH is essential for pIP501 mediated conjugation. Moreover, oligomerization state and topology of TraH in the native membrane were determined providing insights in molecular organization of a Gram-positive T4SS.
抗生素耐药菌株的不断增加所导致的无法治疗的细菌感染，对21世纪的人类医疗保健构成了严重威胁。接合性DNA转移是细菌之间抗生素耐药性和毒力基因传播的最重要机制，由一种称为IV型分泌系统（T4SS）的蛋白质复合物介导。T4SS的核心是一种多蛋白复合物，它作为大分子分泌的通道跨越细菌包膜。我们报告了由接合性革兰氏阳性广宿主范围质粒pIP501编码的转移蛋白TraH的核磁共振结构和功能表征。该结构与革兰氏阴性分泌系统的VirB8蛋白具有惊人的相似性，在分泌机制的支架中发挥着重要作用。考虑到TraM是在pIP501中发现的第一个类似VirB8的蛋白，TraH代表了相应转移操纵子中属于该家族的第二个蛋白。与pIP501野生型（wt）质粒相比，pIP501中无标记的traH缺失导致粪肠球菌中的转移完全丧失，表明TraH对于pIP501介导的接合至关重要。此外，确定了TraH在天然膜中的寡聚化状态和拓扑结构，为革兰氏阳性T4SS的分子组织提供了见解。
