Excursion of a single polypeptide into a protein pore: simple physics, but complicated biology
Excursion of a single polypeptide into a protein pore: simple physics, but complicated biology
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DOI：
10.1007/s00249-008-0309-9
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发表时间：
2008-07-01
影响因子：
2
通讯作者：
Movileanu, Liviu
中科院分区：
文献类型：
--
作者：
Mohammad, M. Mohammad;Movileanu, Liviu
尽管其在分子生物学和实际医学生物技术中的基本和关键重要性，但多肽如何与跨膜蛋白孔相互作用尚未得到全面理解。在这里，我们采用单通道电记录来揭示短多肽和小折叠蛋白质与稳健的β-桶蛋白质孔的相互作用。短多肽的长度类似于25个残基，类似于参与蛋白质输入的带正电荷的靶向前序列。蛋白质由带正电荷的前细胞色素B（2）片段（pB（2））与小核糖核酸酶芽孢杆菌RNA酶（类似于110个残基，Ba）融合组成。单分子实验探索了折叠的pb（2）-Ba蛋白与单个β-桶孔（其中含有带负电荷的静电陷阱）的相互作用，揭示了分子间力网络的复杂性，包括驱动力和静电力。此外，这种相互作用还取决于其他因素，如相互作用多肽的疏水含量、静电陷阱的位置、pb（2）前序列的长度和温度。这种单分子方法与蛋白质设计的相互作用的多肽或孔腔开辟了新的机会，探索在高时间分辨率的多肽孔相互作用。这些未来的研究也有望揭示纳米孔技术用于检测和探索单个多肽的优点和局限性。
Despite its fundamental and critical importance in molecular biology and practical medical biotechnology, how a polypeptide interacts with a transmembrane protein pore is not yet comprehensively understood. Here, we employed single-channel electrical recordings to reveal the interactions of short polypeptides and small folded proteins with a robust beta-barrel protein pore. The short polypeptides were similar to 25 residues in length, resembling positively charged targeting presequences involved in protein import. The proteins were consisted of positively charged pre-cytochrome b(2) fragments (pb(2)) fused to the small ribonuclease barnase (similar to 110 residues, Ba). Single-molecule experiments exploring the interaction of a folded pb(2)-Ba protein with a single beta-barrel pore, which contained negatively charged electrostatic traps, revealed the complexity of a network of intermolecular forces, including driving and electrostatic ones. In addition, the interaction was dependent on other factors, such as the hydrophobic content of the interacting polypeptide, the location of the electrostatic trap, the length of the pb(2) presequence and temperature. This single-molecule approach together with protein design of either the interacting polypeptide or the pore lumen opens new opportunities for the exploration of the polypeptide-pore interaction at high temporal resolution. Such future studies are also expected to unravel the advantages and limitations of the nanopore technique for the detection and exploration of individual polypeptides.