Stability of the proteasome can be regulated allosterically through engagement of its proteolytic active sites
Stability of the proteasome can be regulated allosterically through engagement of its proteolytic active sites
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DOI：
10.1038/nsmb1335
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发表时间：
2007-12-01
影响因子：
16.8
通讯作者：
Finley, Daniel
中科院分区：
文献类型：
--
作者：
Kleijnen, Maurits F.;Roelofs, Jeroen;Finley, Daniel
26 S蛋白酶体全酶由20 S核心颗粒（CP）与19 S调节颗粒（RP）结合形成。CP-RP相互作用是不稳定的，在体内受到调节，但控制这种关联的因素知之甚少。在这里，我们描述了一种体外蛋白酶体重建试验和高分辨率，二维凝胶电泳系统。使用这些技术，我们发现，酵母CP-RP复合物可以包含亚化学计量量的紧密结合，基本上是不可交换的ATP。然而，如果CP-RP复合物被Ecm 29蛋白保留，则该核苷酸被限制用于CP通道的门控。出乎意料的是，蛋白酶体抑制剂在稳定蛋白酶体对抗CP-RP解离方面是有效的。这些数据表明，活性位点的CP与绑定RP通信，尽管他们的空间分离。我们建议，正在进行的蛋白质降解可能会抑制蛋白酶体解体，从而提高蛋白质水解的持续性。
The 26S proteasome holoenzyme is formed by the association of a 20S core particle (CP) with a 19S regulatory particle ( RP). The CP-RP interaction is labile and subject to regulation in vivo, but the factors controlling this association are poorly understood. Here we describe an in vitro proteasome reconstitution assay and a high-resolution, two-dimensional gel electrophoresis system. Using these techniques, we find that a yeast CP-RP complex can contain a substoichiometric amount of tightly bound, essentially non-exchangeable ATP. However, this nucleotide is dispensable for gating of the CP channel, provided that the CP-RP complex is preserved by the Ecm29 protein. Unexpectedly, proteasome inhibitors are potent in stabilizing proteasomes against the dissociation of CP-RP. These data indicate that active sites of the CP communicate with bound RP, despite their spatial separation. We propose that ongoing protein degradation may suppress proteasome disassembly, thereby enhancing the processivity of proteolysis.