Bodrenko Igor V, Wang Jiajun, Salis Samuele, Winterhalter Mathias, Ceccarelli Matteo
Department of Physics, University of Cagliari , S.P. Monserrato-Sestu km 0.700, I-09042, Monserrato, Cagliari, Italy.
Jacobs University , Campus Ring 1, 28759 Bremen, Germany.
ACS Sens. 2017 Aug 25;2(8):1184-1190. doi: 10.1021/acssensors.7b00311. Epub 2017 Jul 28.
To quantify small molecule penetration into and eventually permeation through nanopores, we applied an improved excess-noise analysis of the ion current fluctuation caused by entering molecules. The kinetic parameters of substrate entry and exit are derived from a two-state Markov model, analyzing the substrate concentration dependence of the average ion current and its variance. Including filter corrections allows one to detect the transition rates beyond the cutoff frequency, f, of the instrumental ion-current filter. As an application of the method, we performed an analysis of the single-channel ion current of Meropenem, an antibiotic of the carbapenem family, interacting with OmpF, the major general outer membrane channel of Escherichia coli bacteria. At 40 °C we detected the residence time of Meropenem inside OmpF of about 500 ns-more than 2 orders of magnitude smaller than f and close to the diffusion limit of few hundred nanoseconds. We also have established theoretical limit conditions under which the substrate-induced channel blockages can be detected and suggest that submicrosecond-scale gating kinetic parameters are accessible with existing experimental equipment.
为了量化小分子进入并最终透过纳米孔的情况，我们对由进入分子引起的离子电流波动应用了改进的过量噪声分析。底物进出的动力学参数源自双态马尔可夫模型，通过分析平均离子电流及其方差对底物浓度的依赖性来得出。纳入滤波器校正可以检测超出仪器离子电流滤波器截止频率f的跃迁速率。作为该方法的一个应用，我们对美罗培南（一种碳青霉烯类抗生素）与大肠杆菌主要外膜通道OmpF相互作用的单通道离子电流进行了分析。在40°C时，我们检测到美罗培南在OmpF内的停留时间约为500纳秒，比f小两个数量级以上，且接近几百纳秒的扩散极限。我们还建立了能够检测底物诱导通道阻断的理论极限条件，并表明利用现有实验设备可以获取亚微秒级的门控动力学参数。
