ATP binding to neighbouring subunits and intersubunit allosteric coupling underlie proteasomal ATPase function.
ATP binding to neighbouring subunits and intersubunit allosteric coupling underlie proteasomal ATPase function.
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DOI：
10.1038/ncomms9520
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发表时间：
2015-10-14
影响因子：
16.6
通讯作者：
Smith DM
中科院分区：
文献类型：
--
作者：
Kim YC;Snoberger A;Schupp J;Smith DM
蛋白酶体的主要功能是由一个高度变构的atp酶复合物驱动的。ATP仅与该六聚体中的两个亚基结合，触发底物结合，ATP - 20S结合和20S门打开。然而，目前尚不清楚ATP结合和水解如何在空间和时间上协调这些变构效应，从而驱动底物转运进入20S。在这里，我们使用FRET显示真核生物（RPTs）和古细菌（PAN）的蛋白酶体ATP酶在邻近亚基上具有高亲和力，这补充了26S ATP酶的螺旋阶梯拓扑结构。我们进一步表明，位于亚基界面的PAN中两个保守的精氨酸手指作为一个单一的变构单位一起工作，介导ATP结合的变构效应，而不改变核苷酸结合模式。快速动力学分析还表明，顺序水解机制的环重置可以用ATP的热力学平衡结合来解释。这些数据支持一个模型，即这两个功能不同的变构网络合作将多肽转运到20S进行降解。26S蛋白酶体含有atp酶亚基的六聚体，以atp依赖的方式结合、展开和转运底物。Kim等人使用FRET表明ATP结合优先发生在六聚体的邻近亚基上，并确定了协调易位的两个变构系统。
The primary functions of the proteasome are driven by a highly allosteric ATPase complex. ATP binding to only two subunits in this hexameric complex triggers substrate binding, ATPase–20S association and 20S gate opening. However, it is unclear how ATP binding and hydrolysis spatially and temporally coordinates these allosteric effects to drive substrate translocation into the 20S. Here, we use FRET to show that the proteasomal ATPases from eukaryotes (RPTs) and archaea (PAN) bind ATP with high affinity at neighbouring subunits, which complements the well-established spiral-staircase topology of the 26S ATPases. We further show that two conserved arginine fingers in PAN located at the subunit interface work together as a single allosteric unit to mediate the allosteric effects of ATP binding, without altering the nucleotide-binding pattern. Rapid kinetics analysis also shows that ring resetting of a sequential hydrolysis mechanism can be explained by thermodynamic equilibrium binding of ATP. These data support a model whereby these two functionally distinct allosteric networks cooperate to translocate polypeptides into the 20S for degradation. The 26S proteasome contains a hexamer of ATPase subunits, which binds, unfolds and translocates substrates in an ATP-dependent manner. Kim et al. use FRET to show that ATP binding preferentially occurs at neighbouring subunits of the hexamer, and identify two allosteric systems that coordinate translocation.