Hyde Eva I, Callow Philip, Rajasekar Karthik V, Timmins Peter, Patel Trushar R, Siligardi Giuliano, Hussain Rohanah, White Scott A, Thomas Christopher M, Scott David J
School of Biosciences, University of Birmingham, Birmingham B15 2TT, U.K.
Institut Laue Langevin, 71 avenue des Martyrs, CS 20156, 38042 Grenoble Cedex 9, France.
Biochem J. 2017 Aug 30;474(18):3121-3135. doi: 10.1042/BCJ20170281.
The ParB protein, KorB, from the RK2 plasmid is required for DNA partitioning and transcriptional repression. It acts co-operatively with other proteins, including the repressor KorA. Like many multifunctional proteins, KorB contains regions of intrinsically disordered structure, existing in a large ensemble of interconverting conformations. Using NMR spectroscopy, circular dichroism and small-angle neutron scattering, we studied KorB selectively within its binary complexes with KorA and DNA, and within the ternary KorA/KorB/DNA complex. The bound KorB protein remains disordered with a mobile C-terminal domain and no changes in the secondary structure, but increases in the radius of gyration on complex formation. Comparison of wild-type KorB with an N-terminal deletion mutant allows a model of the ensemble average distances between the domains when bound to DNA. We propose that the positive co-operativity between KorB, KorA and DNA results from conformational restriction of KorB on binding each partner, while maintaining disorder.
来自RK2质粒的ParB蛋白KorB是DNA分配和转录抑制所必需的。它与包括阻遏蛋白KorA在内的其他蛋白质协同作用。像许多多功能蛋白质一样,KorB包含内在无序结构区域,以大量相互转换的构象存在。我们使用核磁共振光谱、圆二色性和小角中子散射技术,在KorB与KorA和DNA的二元复合物以及三元KorA/KorB/DNA复合物中对KorB进行了选择性研究。结合的KorB蛋白仍然无序,其C末端结构域可移动,二级结构没有变化,但在形成复合物时回转半径增加。将野生型KorB与N末端缺失突变体进行比较,可以得到结合DNA时各结构域之间的整体平均距离模型。我们提出,KorB、KorA和DNA之间的正协同作用源于KorB在结合每个伙伴时的构象限制,同时保持无序状态。