Rotation mechanism of Enterococcus hirae V1-ATPase based on asymmetric crystal structures
Rotation mechanism of Enterococcus hirae V1-ATPase based on asymmetric crystal structures
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DOI：
10.1038/nature11778
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发表时间：
2013-01-31
期刊：
影响因子：
64.8
通讯作者：
Murata, Takeshi
中科院分区：
文献类型：
--
作者：
Arai, Satoshi;Saijo, Shinya;Murata, Takeshi
在各种细胞膜系统中，空泡ATP酶（V-ATP酶）作为质子泵发挥作用，参与骨吸收和癌症转移等许多过程，这些膜蛋白是骨质疏松症和癌症的有吸引力的药物靶点（1）。亲水性V-1部分被称为旋转马达，其中中心轴DF复合物利用ATP水解能量在六边形排列的催化A（3）B（3）复合物内旋转，但由于缺乏高分辨率结构信息，分子机制尚未明确。我们先前报道了肠球菌肠溶酶V-1-ATP酶从A（3）B（3）和DF复合物（2，3）的体外表达、纯化和重建。在这里，我们报告的不对称结构的核苷酸自由（2.8埃）和核苷酸结合（3.4埃）的A（3）B（3）复合物，证明构象变化引起的核苷酸结合，表明在右手旋转方向的合作方式的结合顺序。还报道了无核苷酸（2.2埃）和核苷酸结合（2.7埃）的V-1-ATP酶的晶体结构。更紧密包装的核苷酸结合位点似乎是由DF结合诱导的，ATP水解似乎是由保守的精氨酸残基的方法刺激的。据我们所知，这些不对称结构代表了V-1-ATP酶旋转机制的第一个高分辨率视图。
In various cellular membrane systems, vacuolar ATPases (V-ATPases) function as proton pumps, which are involved in many processes such as bone resorption and cancer metastasis, and these membrane proteins represent attractive drug targets for osteoporosis and cancer(1). The hydrophilic V-1 portion is known as a rotary motor, in which a central axis DF complex rotates inside a hexagonally arranged catalytic A(3)B(3) complex using ATP hydrolysis energy, but the molecular mechanism is not well defined owing to a lack of high-resolution structural information. We previously reported on the in vitro expression, purification and reconstitution of Enterococcus hirae V-1-ATPase from the A(3)B(3) and DF complexes(2,3). Here we report the asymmetric structures of the nucleotide-free (2.8 angstrom) and nucleotide-bound (3.4 angstrom) A(3)B(3) complex that demonstrate conformational changes induced by nucleotide binding, suggesting a binding order in the right-handed rotational orientation in a cooperative manner. The crystal structures of the nucleotide-free (2.2 angstrom) and nucleotide-bound (2.7 angstrom) V-1-ATPase are also reported. The more tightly packed nucleotide-binding site seems to be induced by DF binding, and ATP hydrolysis seems to be stimulated by the approach of a conserved arginine residue. To our knowledge, these asymmetric structures represent the first high-resolution view of the rotational mechanism of V-1-ATPase.