Engineering of a novel Ca^<2+>-regulated kinesin molecular motor using a calmodulin dimer linker
Engineering of a novel Ca^<2+>-regulated kinesin molecular motor using a calmodulin dimer linker
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使用钙调蛋白二聚体连接体设计新型 Ca^2 调节的驱动蛋白分子马达
DOI：
10.1016/j.bbrc.2012.05.135
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发表时间：
2012
影响因子：
3.1
通讯作者：
Hideki Shishido
中科院分区：
文献类型：
--
作者：
Kaori Onishi;Jumpei Hamatsu;Dambarudhar Shiba Sanker Hembram;Takehiro Haremaki;Teppei Ikeya;Masaki Mishima;Masahiro Shirakawa and Yutaka Ito;Kodama Y.;Hideki Shishido
激动素-微管系统作为生物芯片中的分子穿梭设备有着巨大的前景。然而，目前的一个障碍是，这样的航天飞机无法对其运动进行“开关”控制。在这里，我们报道了一种新型的分子马达的开发，该马达由加速器和制动系统驱动，使用动蛋白单体和钙调蛋白(CaM)二聚体。将激动素单体K355与CaM靶肽(M13肽)在颈区(K355-M13)的C末端融合。我们还利用CaM突变体(Q3C)、(R86C)或(A147C)以及与半胱氨酸残基反应的交联剂制备了CaM二聚体。在用CaM二聚体诱导K355-M13二聚化后，我们测量了K355-M13的运动能力，发现它可以以钙依赖的方式可逆地调节。我们还发现，K355-M13的运动速度随所使用的CaM二聚体的类型和交联物位置的不同而不同；交联物的长度对运动也有适度的影响。这些结果表明，依赖于钙离子的K355-M13二聚化可以作为一种新型的分子航天器，配备加速器和制动系统，用于生物芯片应用。
The kinesin–microtubule system holds great promise as a molecular shuttle device within biochips. However, one current barrier is that such shuttles do not have “on–off” control of their movement. Here we report the development of a novel molecular motor powered by an accelerator and brake system, using a kinesin monomer and a calmodulin (CaM) dimer. The kinesin monomer, K355, was fused with a CaM target peptide (M13 peptide) at the C-terminal part of the neck region (K355–M13). We also prepared CaM dimers using CaM mutants (Q3C), (R86C), or (A147C) and crosslinkers that react with cysteine residues. Following induction of K355–M13 dimerization with CaM dimers, we measured K355–M13 motility and found that it can be reversibly regulated in a Ca2+-dependent manner. We also found that velocities of K355–M13 varied depending on the type and crosslink position of the CaM dimer used; crosslink length also had a moderate effect on motility. These results suggest Ca2+-dependent dimerization of K355–M13 could be used as a novel molecular shuttle, equipped with an accelerator and brake system, for biochip applications.