An outer membrane protein undergoes enthalpy- and entropy-driven transitions.
An outer membrane protein undergoes enthalpy- and entropy-driven transitions.
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DOI：
10.1021/bi300332z
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发表时间：
2012-07-03
期刊：
影响因子：
2.9
通讯作者：
Movileanu L
中科院分区：
文献类型：
--
作者：
Cheneke BR;Indic M;van den Berg B;Movileanu L
β-桶膜蛋白经常在各种开放亚状态之间波动，但这些转变的性质尚未完全了解。使用温度依赖性、单分子电生理学分析，沿着合理的蛋白质设计，我们表明，铜绿假单胞菌外膜羧酸通道的成员OccK 1具有离散的门控动力学特征，包括焓驱动和熵驱动的电流转变。选择OccK 1来分析这些跃迁，因为它是已知高分辨率晶体结构的单体跨膜β-桶，并显示出三种可区分的、时间可分辨的开放子状态。天然和环缺失OccK 1蛋白质的通道转换的激活熵和熵的显着变化，但在平衡自由能的适度改变，证实了该系统永远不会离开平衡。此外，OccK 1的一些电流波动表明了一个违反直觉的负活化焓，这是由活化熵的显着减少补偿。对OccK 1单通道特性的温度扫描显示，在4°C的最低检测温度下，能量最有利的打开子状态的热诱导开关。因此，这种对电流波动动力学的半定量评估不仅证明了通道门控的复杂性，而且还揭示了β-桶外膜蛋白在不同温度环境下的不同功能特征。
β-barrel membrane proteins often fluctuate among various open sub-states, yet the nature of these transitions is not fully understood. Using temperature-dependent, single-molecule electrophysiology analysis, along with rational protein design, we show that OccK1, a member of the outer membrane carboxylate channel from Pseudomonas aeruginosa, features a discrete gating dynamics comprising of both enthalpy-driven and entropy-driven current transitions. OccK1 was chosen to analyze these transitions, because it is a monomeric transmembrane β-barrel of known high-resolution crystal structure and displays three distinguishable, time-resolvable open sub-states. Native and loop-deletion OccK1 proteins showed substantial changes in the activation enthalpies and entropies of the channel transitions, but modest alterations in the equilibrium free energies, confirming that the system never departs from equilibrium. Moreover, some current fluctuations of OccK1 indicated a counterintuitive, negative activation enthalpy, which was compensated by a significant decrease in the activation entropy. Temperature scanning of the single-channel properties of OccK1 exhibited a thermally-induced switch of the energetically most favorable open sub-state at the lowest examined temperature of 4°C. Therefore, such a semi-quantitative assessment of the current fluctuation dynamics not only demonstrates the complexity of channel gating, but also reveals distinct functional traits of a β-barrel outer membrane protein under different temperature circumstances.
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DOI：
10.1007/s00249-008-0366-0
发表时间：
2008-11-01
影响因子：
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作者：
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通讯作者：
Kleinekathoefer, Ulrich