Aoki Takahiro, Tomishige Michio, Ariga Takayuki
Department of Applied Physics, School of Engineering, the University of Tokyo, Tokyo, 113-8656, Japan.
Biophysics (Nagoya-shi). 2013 Nov 7;9:149-59. doi: 10.2142/biophysics.9.149. eCollection 2013.
Kinesin-1 (conventional kinesin) is a molecular motor that transports various cargo such as endoplasmic reticulum and mitochondria in cells. Its two head domains walk along microtubule by hydrolyzing ATP, while the tail domains at the end of the long stalk bind to the cargo. When a kinesin is not carrying cargo, its motility and ATPase activity is inhibited by direct interactions between the tail and head. However, the mechanism of this tail regulation is not well understood. Here, we apply single molecule fluorescence resonance energy transfer (smFRET) to observe this interaction in stalk-truncated kinesin. We found that kinesin with two tails forms a folding conformation and dissociates from microtubules, whereas kinesin with one tail remains bound to the micro-tubule and is immobile even in the presence of ATP. We further investigated the head-tail interaction as well as head-head coordination on the microtubule at various nucleotide conditions. From these results, we propose a two-step inhibition model for kinesin motility.
驱动蛋白-1（传统驱动蛋白）是一种分子马达，可在细胞中运输各种货物，如内质网和线粒体。它的两个头部结构域通过水解ATP沿着微管移动，而长柄末端的尾部结构域则与货物结合。当驱动蛋白不携带货物时，其运动性和ATP酶活性会受到尾部和头部之间直接相互作用的抑制。然而，这种尾部调节的机制尚未得到很好的理解。在这里，我们应用单分子荧光共振能量转移（smFRET）来观察截短柄部的驱动蛋白中的这种相互作用。我们发现，有两条尾巴的驱动蛋白形成一种折叠构象并从微管上解离，而有一条尾巴的驱动蛋白即使在有ATP的情况下仍与微管结合且不移动。我们进一步研究了在各种核苷酸条件下微管上的头尾相互作用以及头头协同作用。根据这些结果，我们提出了一种驱动蛋白运动性的两步抑制模型。
