Processive movement of single kinesins on crowded microtubules visualized using quantum dots
Processive movement of single kinesins on crowded microtubules visualized using quantum dots
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DOI：
10.1038/sj.emboj.7600937
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发表时间：
2006-01-25
期刊：
影响因子：
11.4
通讯作者：
Surrey, T
中科院分区：
文献类型：
--
作者：
Seitz, A;Surrey, T
动力蛋白-1是一种沿微管运输货物的过程性分子马达。在细胞内，几个马达和微管相关蛋白竞争与微管结合。因此，问题出现了运动蛋白1的进程如何受到微管上拥挤的影响。在这里，我们使用全内反射荧光显微镜对稳态条件下拥挤微管上的单量子点标记的运动蛋白进行体外成像，并测量稳态下微管上的拥挤程度。我们发现微管上的高运动蛋白密度对运动蛋白的流动影响很小。然而，高密度突变的激酶蛋白不能有效地步进，这降低了野生型激酶蛋白的平均速度，而几乎没有改变其加工能力。这表明当遇到障碍时，激酶蛋白在微管上处于强结合状态，直到障碍解除结合，释放结合位点供激酶蛋白进行下一步。一个简单的动力学模型可以定量地解释动力蛋白在两种拥挤条件下的行为。
Kinesin-1 is a processive molecular motor transporting cargo along microtubules. Inside cells, several motors and microtubule-associated proteins compete for binding to microtubules. Therefore, the question arises how processive movement of kinesin-1 is affected by crowding on the microtubule. Here we use total internal reflection fluorescence microscopy to image in vitro the runs of single quantum dot-labelled kinesins on crowded microtubules under steady-state conditions and to measure the degree of crowding on a microtubule at steady-state. We find that the runs of kinesins are little affected by high kinesin densities on a microtubule. However, the presence of high densities of a mutant kinesin that is not able to step efficiently reduces the average speed of wild-type kinesin, while hardly changing its processivity. This indicates that kinesin waits in a strongly bound state on the microtubule when encountering an obstacle until the obstacle unbinds and frees the binding site for kinesin's next step. A simple kinetic model can explain quantitatively the behaviour of kinesin under both crowding conditions.