Quasithermodynamic contributions to the fluctuations of a protein nanopore.
Quasithermodynamic contributions to the fluctuations of a protein nanopore.
复制标题
DOI：
10.1021/cb5008025
复制
发表时间：
2015-03-20
影响因子：
4
通讯作者：
Movileanu, Liviu
中科院分区：
文献类型：
--
作者：
Cheneke, Belete R.;van den Berg, Bert;Movileanu, Liviu
蛋白质经历两个或多个亚态之间的热激活构象波动，但它们的动力学的定量调查一直受到许多因素的挑战，包括各种相互作用的复杂性和动力学，沿着无法在可分辨的时间尺度内检测功能亚态。在这里，我们详细分析了已知高分辨率X射线晶体结构的单体β-桶蛋白纳米孔的电流波动。我们证明了蛋白质纳米孔系统的靶向扰动，以环缺失诱变的形式，伴随着长细胞外环之间静电相互作用的改变，产生了在25 °C下计算的微分活化自由能ΔΔG的适度变化，在热能附近的范围内，但微分活化自由能ΔΔH和熵ΔΔS的实质性和相关的修改。这一发现表明，该蛋白质纳米孔的中央收缩内衬的波动环的包装和灵活性的局部构象重组得到了单通道动力学变化的补充。这些变化反映在补偿温度TC为300 K，活化自由能常数为41 kJ/mol的环伴侣之间的相互作用的熵-熵再转换中。我们还确定，温度对蛋白质纳米孔的平衡门控波动的能量学的影响比其他环境参数大得多，例如水相的离子强度以及所施加的跨膜电位，这可能是由于溶剂化活化过程中的大量变化。将这种方法应用于其他复杂的多状态膜蛋白系统没有根本的限制。因此，这种方法在膜蛋白设计和动力学领域具有重要意义，主要是通过揭示多个明确定义和功能不同的蛋白质通道和孔亚态之间的平衡转换的更好的定量评估。
Proteins undergo thermally activated conformational fluctuations among two or more substates, but a quantitative inquiry on their kinetics is persistently challenged by numerous factors, including the complexity and dynamics of various interactions, along with the inability to detect functional substates within a resolvable time scale. Here, we analyzed in detail the current fluctuations of a monomeric β-barrel protein nanopore of known high-resolution X-ray crystal structure. We demonstrated that targeted perturbations of the protein nanopore system, in the form of loop-deletion mutagenesis, accompanying alterations of electrostatic interactions between long extracellular loops, produced modest changes of the differential activation free energies calculated at 25 °C, ΔΔG⧧, in the range near the thermal energy but substantial and correlated modifications of the differential activation enthalpies, ΔΔH⧧, and entropies, ΔΔS⧧. This finding indicates that the local conformational reorganizations of the packing and flexibility of the fluctuating loops lining the central constriction of this protein nanopore were supplemented by changes in the single-channel kinetics. These changes were reflected in the enthalpy–entropy reconversions of the interactions between the loop partners with a compensating temperature, TC, of ∼300 K, and an activation free energy constant of ∼41 kJ/mol. We also determined that temperature has a much greater effect on the energetics of the equilibrium gating fluctuations of a protein nanopore than other environmental parameters, such as the ionic strength of the aqueous phase as well as the applied transmembrane potential, likely due to ample changes in the solvation activation enthalpies. There is no fundamental limitation for applying this approach to other complex, multistate membrane protein systems. Therefore, this methodology has major implications in the area of membrane protein design and dynamics, primarily by revealing a better quantitative assessment on the equilibrium transitions among multiple well-defined and functionally distinct substates of protein channels and pores.
登录
查看更多内容
DOI：
10.1073/pnas.0711561105
发表时间：
2008-04-29
影响因子：
11.1
作者：
Chen, Min;Khalid, Syma;Bayley, Hagan
通讯作者：
Bayley, Hagan
影响因子：
4.8
作者：
Eren, Elif;Parkin, Jamie;van den Berg, Bert
通讯作者：
van den Berg, Bert
影响因子：
14.8
作者：
Hong, Heedeok;Szabo, Gabor;Tamm, Lukas K.
通讯作者：
Tamm, Lukas K.
影响因子：
5.6
作者：
Basle, Arnaud;Rummel, Gabriele;Schirmer, Tilman
通讯作者：
Schirmer, Tilman
影响因子：
16.8
作者：
Biswas, Shyamasri;Mohammad, M. Mohammad;van den Berg, Bert
通讯作者：
van den Berg, Bert