Tracking single proteins within cells
Tracking single proteins within cells
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DOI：
10.1016/s0006-3495(00)76467-8
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发表时间：
2000-10-01
影响因子：
3.4
通讯作者：
Simon, SM
中科院分区：
文献类型：
--
作者：
Goulian, M;Simon, SM
我们提出了对哺乳动物细胞内的单个蛋白质进行成像和追踪的实验。通过落射荧光显微镜以 71 帧/秒的速度对核质和细胞质中的 R-藻红蛋白 (RPE) 单一蛋白进行成像。我们获得了细胞质中平均 17 帧和细胞核中 16 帧的蛋白质二维轨迹（对应于三维轨迹在焦平面上的投影）。扩散常数由均方位移的线性拟合和每帧的平均位移平方确定。我们发现细胞内 RPE 的扩散常数分布比从甘油溶液中的 RPE、蒙特卡特模拟和简单扩散的理论分布获得的分布更宽。这表明在我们测量的时间尺度上，细胞质和核质中单个 RPE 蛋白的运动不能通过具有独特扩散常数的简单扩散来建模。我们的结果表明，追踪细胞内单个蛋白质的运动是可能的，并且单分子追踪技术可用于探测细胞内大分子的动态。
We present experiments in which single proteins were imaged and tracked within mammalian cells. Single proteins of R-phycoerythrin (RPE) were imaged by epifluorescence microscopy in the nucleoplasm and cytoplasm at 71 frames/s. We acquired two-dimensional trajectories of proteins (corresponding to the projection of three-dimensional trajectories onto the plane of focus) for an average of 17 frames in the cytoplasm and 16 frames in the nucleus. Diffusion constants were determined from linear fits to the mean square displacement and from the mean displacement squared per frame. We find that the distribution of diffusion constants for RPE within cells is broader than the distributions obtained from RPE in a glycerol solution, from a Monte Carte simulation, and from the theoretical distribution for simple diffusion. This suggests that on the time scales of our measurements, the motion of single RPE proteins in the cytoplasm and nucleoplasm cannot be modeled by simple diffusion with a unique diffusion constant. Our results demonstrate that it is possible to follow the motion of single proteins within cells and that the technique of single molecule tracking can be used to probe the dynamics of intracellular macromolecules.