Institut de Pharmacologie et de Biologie Structurale (IPBS), Université de Toulouse, CNRS, UPS, Toulouse, France.
Centre de Biologie Structurale, CNRS, Université de Montpellier, INSERM, 34090, Montpellier, France.
Nat Commun. 2020 Dec 1;11(1):6140. doi: 10.1038/s41467-020-19934-z.
Hydrogen-Deuterium eXchange coupled to Mass Spectrometry (HDX-MS) is now common practice in structural biology. However, it is most of the time applied to rather small oligomeric complexes. Here, we report on the use of HDX-MS to investigate conformational differences between the human standard 20S (std20S) and immuno 20S (i20s) proteasomes alone or in complex with PA28αβ or PA28γ activators. Their solvent accessibility is analyzed through a dedicated bioinformatic pipeline including stringent statistical analysis and 3D visualization. These data confirm the existence of allosteric differences between the std20S and i20S at the surface of the α-ring triggered from inside the catalytic β-ring. Additionally, binding of the PA28 regulators to the 20S proteasomes modify solvent accessibility due to conformational changes of the β-rings. This work is not only a proof-of-concept that HDX-MS can be used to get structural insights on large multi-protein complexes in solution, it also demonstrates that the binding of the std20S or i20S subtype to any of its PA28 activator triggers allosteric changes that are specific to this 20S/PA28 pair.
氢氘交换结合质谱（HDX-MS）现在已成为结构生物学中的常用方法。然而，它通常应用于较小的寡聚复合物。在这里，我们报告了使用 HDX-MS 来研究单独或与 PA28αβ 或 PA28γ 激活剂复合的人标准 20S（std20S）和免疫 20S（i20s）蛋白酶体之间的构象差异。通过包括严格的统计分析和 3D 可视化的专用生物信息学管道分析它们的溶剂可及性。这些数据证实了在催化β环内部从α环表面触发的标准 20S 和 i20S 之间存在变构差异。此外，PA28 调节剂与 20S 蛋白酶体的结合由于β环的构象变化而改变了溶剂可及性。这项工作不仅证明了 HDX-MS 可用于研究溶液中大型多蛋白复合物的结构见解，还证明了 std20S 或 i20S 亚型与任何其 PA28 激活剂的结合都会引发特定于该 20S/PA28 对的变构变化。
