T7 DNA helicase: A molecular motor that processively and unidirectionally translocates along single-stranded DNA
T7 DNA helicase: A molecular motor that processively and unidirectionally translocates along single-stranded DNA
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DOI：
10.1016/s0022-2836(02)00733-7
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发表时间：
2002-08-30
影响因子：
5.6
通讯作者：
Patel, SS
中科院分区：
文献类型：
--
作者：
Kim, DE;Narayan, M;Patel, SS
DNA解旋酶是分子马达，其使用来自NTP水解的能量来驱动双链DNA链分离的过程。在这里，我们测量的易位和能量耦合效率的复制DNA解旋酶从噬菌体T7，是一类解旋酶的成员，组装成环状六聚体。利用真实的时间分析法测定了DNA刺激下T7解旋酶dTTP水解活性的稳态动力学，为T7解旋酶沿着ssDNA单向移位提供了依据.动力学数据的全局拟合提供了在18 degreesC下每六聚体每秒132个碱基的平均易位速率。当沿着ssDNA移位时，T7解旋酶以每秒每六聚体49 dTTP的速率水解dTTP，这表明来自一个dTTP水解的能量驱动T7解旋酶沿着ssDNA的沿着两到三个碱基的单向移动。区分该环解旋酶的特征之一是其持续合成能力，其被确定为0.99996，这表明T7解旋酶在解离之前平均行进约75kb的ssDNA。我们提出T7解旋酶以有效的方式沿沿着ssDNA单向移位的能力在DNA解旋中起着至关重要的作用。(C)2002爱思唯尔科技有限公司。保留所有权利。
DNA helicases are molecular motors that use the energy from NTP hydrolysis to drive the process of duplex DNA strand separation. Here, we measure the translocation and energy coupling efficiency of a replicative DNA helicase from bacteriophage T7 that is a member of a class of helicases that assembles into ring-shaped hexamers. Presteady state kinetics of DNA-stimulated dTTP hydrolysis activity of T7 helicase were measured using a real time assay as a function of ssDNA length, which provided evidence for unidirectional translocation of T7 helicase along ssDNA. Global fitting of the kinetic data provided an average translocation rate of 132 bases per second per hexamer at 18 degreesC. While translocating along ssDNA, T7 helicase hydrolyzes dTTP at a rate of 49 dTTP per second per hexamer, which indicates that the energy from hydrolysis of one dTTP drives unidirectional movement of T7 helicase along two to three bases of ssDNA. One of the features that distinguishes this ring helicase is its processivity, which was determined to be 0.99996, which indicated that T7 helicase travels on an average about 75 kb of ssDNA before dissociating. We propose that the ability of T7 helicase to trans-locate unidirectionally along ssDNA in an efficient manner plays a crucial role in DNA unwinding. (C) 2002 Elsevier Science Ltd. All rights reserved.