D'Heygère François, Schwartz Annie, Coste Franck, Castaing Bertrand, Boudvillain Marc
Centre de Biophysique Moléculaire, Centre National de la Recherche Scientifique (CNRS), Rue Charles Sadron, 45071, Orléans, cedex 2, France.
Methods Mol Biol. 2015;1259:293-311. doi: 10.1007/978-1-4939-2214-7_18.
Transcription termination factor Rho is a ring-shaped, homo-hexamieric RNA translocase that dissociates transcription elongation complexes and transcriptional RNA-DNA duplexes (R-loops) in bacteria. The molecular mechanisms underlying these biological functions have been essentially studied with Rho enzymes from Escherichia coli or close Gram-negative relatives. However, phylo-divergent Rho factors may have distinct properties. Here, we describe methods for the preparation and in vitro characterization (ATPase and helicase activities) of the Rho factor from Mycobacterium tuberculosis, a specimen with uncharacteristic molecular and enzymatic features. These methods set the stage for future studies aimed at better defining the diversity of enzymatic properties of Rho across the bacterial kingdom.
转录终止因子Rho是一种环状的同型六聚体RNA转位酶，它能使细菌中的转录延伸复合物和转录RNA-DNA双链体（R环）解离。这些生物学功能背后的分子机制主要是通过对来自大肠杆菌或亲缘关系较近的革兰氏阴性菌的Rho酶进行研究的。然而，系统发育上有差异的Rho因子可能具有不同的特性。在这里，我们描述了结核分枝杆菌Rho因子的制备方法及其体外特性表征（ATP酶和螺旋酶活性），结核分枝杆菌是一种具有非典型分子和酶学特征的样本。这些方法为未来旨在更好地界定细菌界Rho酶特性多样性的研究奠定了基础。
