Meyhöfer E, Howard J
Department of Physiology and Biophysics, University of Washington, Seattle 98195.
Proc Natl Acad Sci U S A. 1995 Jan 17;92(2):574-8. doi: 10.1073/pnas.92.2.574.
To probe the mechanism by which the motor protein kinesin moves along microtubules, we have developed a highly sensitive technique for measuring the force exerted by a single motor molecule. In this technique, one end of a microtubule is attached to the tip of a flexible glass fiber of calibrated stiffness. The other end of the microtubule makes contact with a surface sparsely coated with kinesin. By imaging the tip of the glass fiber on a photodiode detector, displacement of the microtubule by kinesin through as little as 1 nm can be detected and forces as small as 1 pN resolved. Using this force-fiber apparatus we have characterized the mechanical output of this molecular motor. The speed at which a molecule of kinesin moved along the surface of a microtubule decreased linearly as the elastic force was increased. The force required to stop a single kinesin molecule was 5.4 +/- 1.0 pN (mean +/- SD; n = 16), independent of the stiffness of the fiber, the damping from the fluid, and whether the ATP concentration was high or low.
为了探究驱动蛋白这种马达蛋白沿着微管移动的机制，我们开发了一种高度灵敏的技术来测量单个马达分子施加的力。在这项技术中，微管的一端连接到一根具有校准刚度的柔性玻璃纤维的尖端。微管的另一端与稀疏涂有驱动蛋白的表面接触。通过在光电二极管探测器上对玻璃纤维的尖端进行成像，可以检测到驱动蛋白使微管产生的低至1纳米的位移，并分辨出小至1皮牛顿的力。使用这种力纤维装置，我们已经对这种分子马达的机械输出进行了表征。随着弹力增加，驱动蛋白分子沿着微管表面移动的速度呈线性下降。使单个驱动蛋白分子停止移动所需的力为5.4±1.0皮牛顿（平均值±标准差；n = 16），与纤维的刚度、流体的阻尼以及ATP浓度是高还是低无关。
