Mackey A T, Gilbert S P
Department of Biological Sciences, 518 Langley Hall, University of Pittsburgh, PA 15260, USA.
Biochemistry. 2000 Feb 15;39(6):1346-55. doi: 10.1021/bi991918+.
Ncd is a minus-end-directed microtubule motor and a member of the kinesin superfamily. The Ncd dimer contains two motor domains, and cooperative interactions between the heads influence the interactions of each respective motor domain with the microtubule. The approach we have taken to understand the cooperativity between the two motor domains is to analyze the ATPase cycle of dimeric MC1 and monomeric MC6. The steps in the ATPase cycle where cooperativity occurs can be identified by comparing the two mechanisms. The rate-limiting step in the MC6 mechanism is ADP release at 3.4 s(-)(1). The observed rate constant for ATP-induced dissociation from the microtubule is 14 s(-)(1). However, the relative amplitude associated with MC6 dissociation is extremely small in comparison to the amplitude associated with dimeric MC1 dissociation kinetics. The amplitude data indicate that monomeric MC6 does not detach from the microtubule during the initial turnovers of ATP, and ATP hydrolysis is uncoupled from movement. The results show that cooperative interactions between the motor domains of the dimer are required for ATP-dependent dissociation; therefore, one function of the partner motor domain may be to weaken the interaction of the adjacent head with the microtubule.
Ncd是一种向微管负端移动的分子马达，属于驱动蛋白超家族成员。Ncd二聚体包含两个马达结构域，头部之间的协同相互作用影响每个马达结构域与微管的相互作用。我们用于理解两个马达结构域之间协同性的方法是分析二聚体MC1和单体MC6的ATP酶循环。通过比较这两种机制，可以确定ATP酶循环中发生协同作用的步骤。MC6机制中的限速步骤是ADP以3.4 s⁻¹的速率释放。观察到的ATP诱导从微管解离的速率常数为14 s⁻¹。然而，与二聚体MC1解离动力学相关的幅度相比，与MC6解离相关的相对幅度极小。幅度数据表明，在ATP的初始周转过程中，单体MC6不会从微管上脱离，并且ATP水解与运动解偶联。结果表明，二聚体的马达结构域之间的协同相互作用是ATP依赖性解离所必需的；因此，伙伴马达结构域的一个功能可能是削弱相邻头部与微管的相互作用。
