Topological studies on the twin-arginine translocase component TatC
Topological studies on the twin-arginine translocase component TatC
复制标题
DOI：
10.1016/j.femsle.2004.03.048
复制
发表时间：
2004-05-15
影响因子：
2.1
通讯作者：
Brüser, T
中科院分区：
文献类型：
--
作者：
Behrendt, J;Standar, K;Brüser, T
双精氨酸易位（Tat）系统可以将折叠的蛋白质跨生物膜易位。在大肠杆菌已知的 Tat 系统成分中，TatC 是唯一具有多个跨膜结构域的蛋白质。 TatC 对于易位子与 Tat 底物的相互作用非常重要。因此，了解其膜拓扑结构对于理解底物结合和易位子功能至关重要。最近，基于活性 PhoA 报告基因与 TatC 的第二个预测细胞质环的融合，提出了具有四个跨膜结构域的拓扑，这对六个跨膜结构域的计算机预测提出了质疑。在这里，我们报告了 TatC 与拓扑标记酶 PhoA 和 LacZ 的翻译融合研究，为六跨膜结构域拓扑提供了有力的证据。发现第四个跨膜结构域的停止转移能力受到随后的细胞质结构域的强烈影响。细胞质结构域的 PhoA 融合位点处接头序列的存在诱导了 PhoA 泄漏。在一种测试的融合体（S185-PhoA）的情况下，由于第四跨膜结构域（E170）中心的负电荷，停止转移效率已经很低。结果指出了细胞质环对于稳定终止转移序列的重要性，并撤销了 TatC 仅四个跨膜结构域的证据。 (C) 2004 年欧洲微生物学会联合会。由 Elsevier B.V. 出版。保留所有权利。
The twin-arginine translocation (Tat) system can translocate folded proteins across biological membranes. Among the known Tat-system components in Escherichia coli, TatC is the only protein with multiple trans-membrane domains. TatC is important for translocon interactions with Tat Substrates. The knowledge of its membrane topology is therefore crucial for the understanding of substrate binding and translocon function. Recently, based on active PhoA reporter fusions to the second predicted cytoplasmic loop of TatC, a topology with four trans-membrane domains has been suggested, calling in silico predictions of six trans-membrane domains into question. Here we report studies with translational fusions of TatC to the topological marker enzymes PhoA and LacZ which provide strong evidence for a six-trans-membrane domain topology. The stop transfer capacity of the fourth trans-membrane domain was found to be strongly influenced by the succeeding cytoplasmic domain. The presence of linker sequences at PhoA-fusion sites of the cytoplasmic domain induced PhoA leakage. In the case of one tested fusion (S185-PhoA), the stop-transfer efficiency was already low due to the negative charge in the center Of the fourth trans-membrane domain (E170). The results point to the importance of cytoplasmic loops for the stabilization of stop-transfer sequences and revoke evidence for only four trans-membrane domains of TatC. (C) 2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.