Myosin VI steps via a hand-over-hand mechanism with its lever arm undergoing fluctuations when attached to actin
Myosin VI steps via a hand-over-hand mechanism with its lever arm undergoing fluctuations when attached to actin
复制标题
DOI：
10.1074/jbc.c400252200
复制
发表时间：
2004-09-03
影响因子：
4.8
通讯作者：
Sweeney, HL
中科院分区：
文献类型：
--
作者：
Yildiz, A;Park, H;Sweeney, HL
肌凝蛋白VI是一种反向肌凝蛋白马达，作为二聚体，在肌动蛋白上以每一步平均质量中心移动约30纳米的速度前进。我们在肌球蛋白VI的运动结构域或位于其假定的杠杆臂上的两个钙调素（CaMs）的远端用单个荧光团标记。采用FIONA（一纳米精度的荧光成像）技术，观察步长，标准偏差< 1.5 nm，时间分辨率0.5 s，观察时间为分钟。无论探针的位置如何，标记头部的平均步长都接近于60纳米，这有力地支持了一种手拉手运动模型，并排除了独特的肌球蛋白VI插入物分离的模型。然而，CaM探针在平均位置附近显示出较大的空间波动（存在ATP但不存在ADP或没有核苷酸），而运动结构域探针则没有。这支持了肌凝蛋白VI运动模型，其中杠杆臂要么从运动结构域机械地解耦，要么在肌动蛋白上的部分运动周期中进行可逆异构化。
Myosin VI is a reverse direction myosin motor that, as a dimer, moves processively on actin with an average center-of-mass movement of similar to 30 nm for each step. We labeled myosin VI with a single fluorophore on either its motor domain or on the distal of two calmodulins (CaMs) located on its putative lever arm. Using a technique called FIONA ( fluorescence imaging with one nanometer accuracy), step size was observed with a standard deviation of < 1.5 nm, with 0.5-s temporal resolution, and observation times of minutes. Irrespective of probe position, the average step size of a labeled head was similar to 60 nm, strongly supporting a hand-over-hand model of motility and ruling out models in which the unique myosin VI insert comes apart. However, the CaM probe displayed large spatial fluctuations ( presence of ATP but not ADP or no nucleotide) around the mean position, whereas the motor domain probe did not. This supports a model of myosin VI motility in which the lever arm is either mechanically uncoupled from the motor domain or is undergoing reversible isomerization for part of its motile cycle on actin.