细菌IV型分泌系统(T4SS)是一个多功能的大分子易位基因家族,共同能够招募不同的DNA和蛋白质底物,并将它们传递给广泛的细胞类型。目前,对于T4SS如何识别底物库并与特定靶细胞形成生产性接触的了解甚少。尽管T4SS由许多保守的亚基组成,并采用某些保守的结构特征,它们还显示出相当大的组成和结构多样性。这里,我们探索了T4SS的功能多功能性的结构基础,通过系统的缺失和两个共轭系统之间的亚基交换由远缘相关的IncF质粒编码,pED208和F。我们在编码的T4SS中确定了几个具有内在灵活性的区域,嵌合机器的部分或全部功能证明了这一点。交换VirD4样TraDIV型偶联蛋白(T4CPs)产生功能嵌合体,指示底物-TraD和TraD-T4SS界面处的松弛特异性。通过突变分析,我们进一步描绘了TraDT4CPs的结构域,这些结构域有助于募集同源和异源DNA底物。值得注意的是,包括外膜核心复合物的组分交换,一些F特异性亚基,或在没有可检测的菌毛产生的情况下,TraA菌毛蛋白支持DNA转移。在NCBI数据库中测序的肠细菌物种中,我们鉴定了许多含有两个或更多个F-样质粒的菌株和许多缺乏自身转移所需的一个或多个T4SS组分的F质粒。我们证实携带共同居住的宿主细胞,pED208和F精心制作的嵌合T4SS的非自身传播变体,两种质粒的传播都证明了这一点。我们建议T4SS可塑性能够轻松组装功能嵌合体,结构层面的这种内在灵活性可以解释这个超家族在进化过程中的功能多样化,在更直接的时间尺度上,转移缺陷型MGE在自然界中的增殖。
Bacterial type IV secretion systems (T4SSs) are a versatile family of macromolecular translocators, collectively able to recruit diverse DNA and protein substrates and deliver them to a wide range of cell types. Presently, there is little understanding of how T4SSs recognize substrate repertoires and form productive contacts with specific target cells. Although T4SSs are composed of a number of conserved subunits and adopt certain conserved structural features, they also display considerable compositional and structural diversity. Here, we explored the structural bases underlying the functional versatility of T4SSs through systematic deletion and subunit swapping between two conjugation systems encoded by the distantly-related IncF plasmids, pED208 and F. We identified several regions of intrinsic flexibility among the encoded T4SSs, as evidenced by partial or complete functionality of chimeric machines. Swapping of VirD4-like TraD type IV coupling proteins (T4CPs) yielded functional chimeras, indicative of relaxed specificity at the substrate-TraD and TraD-T4SS interfaces. Through mutational analyses, we further delineated domains of the TraD T4CPs contributing to recruitment of cognate vs heterologous DNA substrates. Remarkably, swaps of components comprising the outer membrane core complexes, a few F-specific subunits, or the TraA pilins supported DNA transfer in the absence of detectable pilus production. Among sequenced enterobacterial species in the NCBI database, we identified many strains that harbor two or more F-like plasmids and many F plasmids lacking one or more T4SS components required for self-transfer. We confirmed that host cells carrying co-resident, non-selftransmissible variants of pED208 and F elaborate chimeric T4SSs, as evidenced by transmission of both plasmids. We propose that T4SS plasticity enables the facile assembly of functional chimeras, and this intrinsic flexibility at the structural level can account for functional diversification of this superfamily over evolutionary time and, on a more immediate time-scale, to proliferation of transfer-defective MGEs in nature.