Solution structure of the E. coli TolA C-terminal domain reveals conformational changes upon binding to the phage g3p N-terminal domain
Solution structure of the E. coli TolA C-terminal domain reveals conformational changes upon binding to the phage g3p N-terminal domain
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DOI：
10.1016/j.jmb.2004.12.028
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发表时间：
2005-03-04
影响因子：
5.6
通讯作者：
Blanchard, L
中科院分区：
文献类型：
--
作者：
Deprez, C;Lloubès, R;Blanchard, L
大肠杆菌的Tol-Pal系统是位于细胞膜内的大分子复合物。它参与维持外膜的完整性，并且是摄取两种不同类型的大分子所必需的，即细菌毒素（大肠杆菌素）和丝状噬菌体的 DNA。 TolA 蛋白在这些输入机制中发挥着核心作用。其 C 端结构域 (TolAIII) 通过与大肠菌素的 N 端结构域和噬菌体小外壳基因 3 蛋白 (g3pN1) 的 N 端结构域直接相互作用而参与易位步骤。之前已经观察到 TolAIII 的极端行为，因为 TolAIII 的结构在与不同的成孔大肠杆菌素结合后要么不受影响，要么采用无序构象。在这里，我们通过异核核磁共振波谱解析了游离 TolAIII 的三维结构，并将其与与 g3pN1 结合的 TolAIII 的晶体结构进行比较，以研究 g3pN1 对 TolAIII 三级结构的影响。 g3pN1 结合的 TolAIII 的主链 H-1、N-15 和 C-13 共振也被指定并用于将游离 TolAIII 的溶液结构叠加在 g3pN1-TolAIII 融合蛋白的晶体结构上。这使我们能够追踪 TolAIII 结合后的构象变化。虽然游离 TolAIII 的整体折叠与 g3pN1 结合的 TolAIII 的整体折叠基本相同，但确实发生了二级结构的转变。因此，TolAIII 在体内也与 Pal 和 TolB 相互作用，能够在与各种配偶体结合后调整其构象。讨论了蛋白质结合机制的可能模型，以解释 TolAIII 这种迄今为止未观察到的行为。 (C) 2004 Elsevier Ltd. 保留所有权利。
The Tol-Pal system of Escherichia coli is a macromolecular complex located in the cell envelope. It is involved in maintaining the integrity of the outer membrane and is required for the uptake of two different types of macromolecules, which are bacteriotoxins (colicins) and DNA of filamentous bacteriophages. The TolA protein plays a central role in these import mechanisms. Its C-terminal domain (TolAIII) is involved in the translocation step via direct interaction with the N-terminal domain of colicins and the N-terminal domain of the phage minor coat gene 3 protein (g3pN1). Extreme behaviours of TolAIII have been previously observed, since the structure of TolAIII either remained unaffected or adopted disordered conformation upon binding to different pore-forming colicins. Here, we have solved the 3D structure of free TolAIII by heteronuclear NMR spectroscopy and compared it to the crystal structure of TolAIII bound to g3pN1 in order to study the effect of g3pN1 on the tertiary structure of TolAIII. Backbone H-1, N-15 and C-13 resonances of the g3pN1-bound TolAIII were also assigned and used to superimpose the solution structure of free TolAIII on the crystal structure of the g3pN1-TolAIII fusion protein. This allowed us to track conformational changes of TolAIII upon binding. While the global fold of free TolAIII is mainly identical to that of g3pN1-bound TolAIII, shift of secondary structures does occur. Thus, TolAIII, which interacts also in vivo with Pal and TolB, is able to adapt its conformation upon binding to various partners. Possible models for protein binding mechanisms are discussed to explain this so-far unobserved behaviour of TolAIII. (C) 2004 Elsevier Ltd. All rights reserved.