Prokaryotic ubiquitin-like protein pup is intrinsically disordered.
Prokaryotic ubiquitin-like protein pup is intrinsically disordered.
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DOI：
10.1016/j.jmb.2009.07.018
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发表时间：
2009-09-11
影响因子：
5.6
通讯作者：
Walters, Kylie J.
中科院分区：
文献类型：
--
作者：
Chen, Xiang;Solomon, William C.;Kang, Yang;Cerda-Maira, Francisca;Darwin, K. Heran;Walters, Kylie J.
原核泛素样蛋白Pup通过与Mpa（一种被认为与20 S催化亚基相邻的ATP酶）相互作用，靶向结核分枝杆菌蛋白酶体降解的底物。在真核生物中，泛素被组装成一种聚合物，类似地发出蛋白酶体降解的信号，它采用了一种稳定而紧凑的结构折叠，这种结构折叠适合于其他蛋白质，以实现不同的生物学功能。我们使用NMR光谱来证明，与泛素不同，64个氨基酸的蛋白质Pup本质上是无序的，在C-末端区域具有小的螺旋倾向。我们发现Pup：Mpa相互作用涉及跨越S21-K61的广泛接触表面，并且结合在NMR时间尺度上处于“缓慢”交换状态，从而证明比大多数泛素：泛素受体对更高的亲和力。有趣的是，在滴定实验期间，可观察到中间Pup物质，表明结合后形成一种或多种瞬态。此外，Mpa选择了一种构型，用于游离蛋白质中进行化学交换的区域。这些发现为Pup作为降解信号的功能作用提供了机制性见解。
The prokaryotic ubiquitin-like protein Pup targets substrates for degradation by the Mycobacterium tuberculosis proteasome through its interaction with Mpa, an ATPase that is thought to abut the 20S catalytic subunit. Ubiquitin, which is assembled into a polymer to similarly signal for proteasomal degradation in eukaryotes, adopts a stable and compact structural fold that is adapted into other proteins for diverse biological functions. We used NMR spectroscopy to demonstrate that unlike ubiquitin, the 64 amino acid protein Pup is intrinsically disordered with small helical propensity in the C-terminal region. We found that the Pup:Mpa interaction involves an extensive contact surface that spans S21–K61 and that the binding is in the “slow” exchange regime on the NMR time scale, thus demonstrating higher affinity than most ubiquitin:ubiquitin receptor pairs. Interestingly, during the titration experiment, intermediate Pup species were observable, suggesting the formation of one or more transient state(s) upon binding. Moreover, Mpa selected one configuration for a region undergoing chemical exchange in the free protein. These findings provide mechanistic insights into Pup’s functional role as a degradation signal.
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