Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, and the Zernike Institute for Advanced Materials, University of Groningen, Groningen, The Netherlands.
AMOLF, Science Park 104, 1098 XG, Amsterdam, The Netherlands.
Sci Rep. 2021 Jan 14;11(1):1433. doi: 10.1038/s41598-021-81081-2.
In bacteria, the SecA ATPase provides the driving force for protein secretion via the SecYEG translocon. While the dynamic interplay between SecA and SecYEG in translocation is widely appreciated, it is not clear how SecA associates with the translocon in the crowded cellular environment. We use super-resolution microscopy to directly visualize the dynamics of SecA in Escherichia coli at the single-molecule level. We find that SecA is predominantly associated with and evenly distributed along the cytoplasmic membrane as a homodimer, with only a minor cytosolic fraction. SecA moves along the cell membrane as three distinct but interconvertible diffusional populations: (1) A state loosely associated with the membrane, (2) an integral membrane form, and (3) a temporarily immobile form. Disruption of the proton-motive-force, which is essential for protein secretion, re-localizes a significant portion of SecA to the cytoplasm and results in the transient location of SecA at specific locations at the membrane. The data support a model in which SecA diffuses along the membrane surface to gain access to the SecYEG translocon.
在细菌中，SecA ATP 酶通过 SecYEG 转运蛋白提供蛋白质分泌的驱动力。尽管 SecA 和 SecYEG 在易位过程中的动态相互作用得到了广泛的认可，但 SecA 如何在拥挤的细胞环境中与转运蛋白结合尚不清楚。我们使用超分辨率显微镜在大肠杆菌中单分子水平上直接观察 SecA 的动力学。我们发现 SecA 主要作为同源二聚体与细胞质膜结合并均匀分布，只有一小部分存在于细胞质中。SecA 沿细胞膜移动，有三种不同但可相互转化的扩散种群：（1）与膜松散结合的状态，（2）整合的膜形式，（3）暂时固定的形式。质子动力势的破坏对蛋白质分泌至关重要，它会将 SecA 的很大一部分重新定位到细胞质中，并导致 SecA 在膜上的特定位置暂时出现。这些数据支持了一种模型，即 SecA 沿膜表面扩散以接近 SecYEG 转运蛋白。
