Characterization of the elongasome core PBP2: MreC complex of Helicobacter pylori
Characterization of the elongasome core PBP2: MreC complex of Helicobacter pylori
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DOI：
10.1111/j.1365-2958.2011.07791.x
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发表时间：
2011-10-01
影响因子：
3.6
通讯作者：
Boneca, Ivo G.
中科院分区：
文献类型：
--
作者：
El Ghachi, Meriem;Matteie, Pierre-Jean;Boneca, Ivo G.
细菌细胞形状的定义是一个复杂的过程，需要复杂的大分子机器的多个组件的参与。我们的目的是表征参与细胞形状的螺旋杆菌幽门螺杆菌的决定因素。使用酵母双杂交筛选与关键细胞延伸蛋白PBP 2作为诱饵，我们确定了PBP 2和MreC之间的相互作用。这种相互作用所需的MreC的最小区域范围为氨基酸116至226。使用重组蛋白，我们通过亲和色谱和尺寸排阻色谱以及表面等离子体共振显示PBP 2和MreC形成稳定的复合物。在体内，这两种蛋白质显示出相似的空间定位和它们的复合物具有明显的1：1的化学计量比，这些结果在体外通过分析超离心和化学交联得到证实。PBP 2：MreC复合物的小角X射线散射分析表明，MreC直接与PBP 2的C-末端区域相互作用。PBP 2或MreC的耗尽导致转变为失去活力的球形细胞。最后，MreC与PBP 2在周质空间中的最小相互作用结构域的反式特异性表达导致细胞变圆，表明体内PBP 2/MreC复合物的形成对细胞形态是必需的。
The definition of bacterial cell shape is a complex process requiring the participation of multiple components of an intricate macromolecular machinery. We aimed at characterizing the determinants involved in cell shape of the helical bacterium Helicobacter pylori. Using a yeast two-hybrid screen with the key cell elongation protein PBP2 as bait, we identified an interaction between PBP2 and MreC. The minimal region of MreC required for this interaction ranges from amino acids 116 to 226. Using recombinant proteins, we showed by affinity and size exclusion chromatographies and surface plasmon resonance that PBP2 and MreC form a stable complex. In vivo, the two proteins display a similar spatial localization and their complex has an apparent 1:1 stoichiometry; these results were confirmed in vitro by analytical ultracentrifugation and chemical cross-linking. Small angle X-ray scattering analyses of the PBP2 : MreC complex suggest that MreC interacts directly with the C-terminal region of PBP2. Depletion of either PBP2 or MreC leads to transition into spherical cells that lose viability. Finally, the specific expression in trans of the minimal interacting domain of MreC with PBP2 in the periplasmic space leads to cell rounding, suggesting that the PBP2/MreC complex formation in vivo is essential for cell morphology.