STRUCTURE AND MOLECULAR-WEIGHT OF THE DYNEIN ATPASE
STRUCTURE AND MOLECULAR-WEIGHT OF THE DYNEIN ATPASE
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DOI：
10.1083/jcb.96.3.669
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发表时间：
1983-01-01
影响因子：
7.8
通讯作者：
WALL, JS
中科院分区：
文献类型：
--
作者：
JOHNSON, KA;WALL, JS
通过扫描透射电子显微镜检查动力蛋白。将来自四膜虫纤毛的 30S 动力蛋白样品涂在 C 胶片上，然后将其冷冻干燥并作为未染色、未固定的样本进行检查，或者用硫酸双氧铀负染色。动力蛋白分子的全新图像被揭示，显示 3 个球状头通过 3 条独立的链连接到一个公共碱基。在这些头中，有 2 个看起来相同，直径为 10 nm；第三个头稍大一些（约 12 nm）。颗粒的总长度为35nm。基于未染色颗粒的电子散射强度积分的质量分析给出了 1.95 (.+-. 0.24) 兆道尔顿的分子量。使用动力蛋白修饰的牛脑微管进行每单位长度的质量分析，在动力蛋白显示 24 nm 线性重复的条件下，7 个动力蛋白分子围绕由 14 根原丝组成的微管。未修饰的微管每单位长度的MW为21,000±。 1900 道尔顿/.ANG。相比之下，值为 84,400 .+-。 2200 道尔顿/.ANG。对于完全装饰的微管。这些数据给出了 2.17 (±0.14) 兆道尔顿/动力蛋白分子的 MW，与对分离颗粒的测量结果一致。对在孤立颗粒中观察到的各个球状头进行的质量分析给出了平均值为 416 ± 的 MW 分布。 76 千道尔顿 [kd]。这些数据也可以被视为 2 个头部群体的总和，其中 2/3 的头部位于 .apprx。 400 kd 和 1/3 at .apprx。 550 kd，尽管需要更精确的数据来自信地区分 2 类头部。分析动力蛋白-微管复合物的质量作为距颗粒中线距离的函数，以区分动力蛋白分子的哪一端与微管结合。预测的质量分布与 3 个动力蛋白头朝向微管的模型一致，但显然与相反方向不一致。 3 个球状头可能形成该异源动力蛋白微管系统中的 ATP 敏感位点。动力蛋白分子的根状碱基可能形成纤毛和鞭毛外双联体 A 亚纤维的结构附着位点。根据这些新结果讨论了动力蛋白的结构和功能。
Dynein was examined by scanning transmission electron microscopy. Samples of 30S dynein from Tetrahymena cilia were applied to C films and either were freeze-dried and examined as unstained, unfixed specimens or were negatively stained with uranyl sulfate. A totally new image of the dynein molecule was revealed showing 3 globular heads connected by 3 separate strands to a common base. Of the heads, 2 appeared to be identical and exhibited a diameter of 10 nm; the 3rd head was somewhat larger (.apprx. 12 nm). The overall length of the particle was 35 nm. Mass analysis, based on the integration of electron scattering intensities unstained particles, gave a MW of 1.95 (.+-. 0.24) megadaltons. Mass per unit length analysis was performed using bovine brain microtubules decorated with dynein under conditions where the dynein shows a linear repeat of 24 nm with 7 dynein molecules surrounding a microtubule made up of 14 protofilaments. Undecorated microtubules gave a MW per unit length of 21,000 .+-. 1900 daltons/.ANG. compared to a value of 84,400 .+-. 2200 daltons/.ANG. for the fully decorated microtubules. These data gave a MW of 2.17 (.+-. 0.14) megadaltons/dynein molecule, in agreement with measurements on the isolated particles. Mass analysis of individual globular heads observed in isolated particles gave a MW distribution with a mean of 416 .+-. 76 kdaltons [kd]. These data could also be viewed as the sum of 2 populations of heads with 2/3 of the heads at .apprx. 400 kd and 1/3 at .apprx. 550 kd, although more precise data will be required to distinguish 2 classes of heads with confidence. The mass of the dynein-microtubule complex as a function of distance from the midline of the particle was analyzed to distinguish which end of the dynein molecule was bound to the microtubule. The projected mass distribution was consistent with a model where the 3 dynein heads were oriented toward the microtubule and clearly not consistent with the opposite orientation. The 3 globular heads probably form the ATP-sensitive site in this heterologous dynein-microtubule system. The rootlike base of the dynein molecule probably forms the structural attachment site to the A-subfiber of the outer doublet in cilia and flagella. The structure and function of the dynein are discussed in terms of these new results.