Probing High Permeability of Nuclear Pore Complexes by Scanning Electrochemical Microscopy: Ca2+ Effects on Transport Barriers.
Probing High Permeability of Nuclear Pore Complexes by Scanning Electrochemical Microscopy: Ca2+ Effects on Transport Barriers.
复制标题
DOI：
10.1021/acs.analchem.9b00796
复制
发表时间：
2019-03
影响因子：
7.4
通讯作者：
Pavithra Pathirathna;Ryan J. Balla;Dylan T. Jantz;Niraja Kurapati;Erin R Gramm;Kevin C. Leonard;S. Amemiya
中科院分区：
文献类型：
--
作者：
Pavithra Pathirathna;Ryan J. Balla;Dylan T. Jantz;Niraja Kurapati;Erin R Gramm;Kevin C. Leonard;S. Amemiya
核孔复合物（NPC）在真核细胞的细胞核和细胞质之间的分子运输中起着重要的生物学和生物医学作用。然而，从化学角度来看，这种生物纳米孔是如何通过富含疏水苯丙氨酸和甘氨酸与带电氨基酸混合的高度无序重复的运输屏障，选择性和有效地运输各种物质，包括小分子、蛋白质和rna的，目前还没有得到很好的理解。在这里，我们使用扫描电化学显微镜来成像和测量npc对小氧化还原分子的高渗透率。有效介质理论表明，测得的渗透率是由探针分子通过充满水的纳米孔的扩散易位控制的，它们分别没有来自疏水区或带电区运输障碍的空间或静电障碍。然而，低毫摩尔浓度的Ca2+降低了NPCs的渗透性，Ca2+可以与运输屏障的阴离子区域相互作用，改变其在纳米孔内的空间分布。我们使用原子力显微镜证实，在高Ca2+水平下，npc的运输屏障主要是凹陷的（~ 80%）或纠缠的（~ 20%），而在亚微摩尔的生理Ca2+水平下，npc的运输屏障主要是纠缠的（~ 50%）、凹陷的（~ 25%）和“堵塞的”（~ 25%）构象。我们提出了一个同步Ca2+对npc构象和通透性影响的模型，其中运输屏障被粘滞以降低通透性。值得注意的是，这一结果支持了一个假设，即运输屏障的功能结构不仅由它们的疏水区域维持，而且由带电区域维持。
The nuclear pore complex (NPC) solely mediates molecular transport between the nucleus and cytoplasm of a eukaryotic cell to play important biological and biomedical roles. However, it is not well-understood chemically how this biological nanopore selectively and efficiently transports various substances, including small molecules, proteins, and RNAs by using transport barriers that are rich in highly disordered repeats of hydrophobic phenylalanine and glycine intermingled with charged amino acids. Herein, we employ scanning electrochemical microscopy to image and measure the high permeability of NPCs to small redox molecules. The effective medium theory demonstrates that the measured permeability is controlled by diffusional translocation of probe molecules through water-filled nanopores without steric or electrostatic hindrance from hydrophobic or charged regions of transport barriers, respectively. However, the permeability of NPCs is reduced by a low millimolar concentration of Ca2+, which can interact with anionic regions of transport barriers to alter their spatial distributions within the nanopore. We employ atomic force microscopy to confirm that transport barriers of NPCs are dominantly recessed (∼80%) or entangled (∼20%) at the high Ca2+ level in contrast to authentic populations of entangled (∼50%), recessed (∼25%), and "plugged" (∼25%) conformations at a physiological Ca2+ level of submicromolar. We propose a model for synchronized Ca2+ effects on the conformation and permeability of NPCs, where transport barriers are viscosified to lower permeability. Significantly, this result supports a hypothesis that the functional structure of transport barriers is maintained not only by their hydrophobic regions, but also by charged regions.