Hassan Bakar, Chandravanshi Monika, Ng Martin Y, Negi Hitendra, Wilson Brice A P, Walters Kylie J
Protein Processing Section, Center for Structural Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD, USA.
Molecular Targets Program, Center for Cancer Research, National Cancer Institute, Frederick, MD, USA.
Nat Commun. 2025 Jul 1;16(1):5669. doi: 10.1038/s41467-025-60843-w.
A pleckstrin-like receptor for ubiquitin (Pru) domain in hRpn13 binds ubiquitin and proteasome subunit hRpn2. Here, we report a crystal structure of Pru bound to amino acids at the extreme N-terminus (ENT) of recombinant hRpn13. ENT adopts a U shape with native sequence along one side where M1 is buried in a Pru W108-centered pocket, and non-native sequence along the other with main chain hydrogen bonding to a neighboring Pru of the crystal lattice. These ENT:Pru interactions are stable in molecular dynamics simulations even with inclusion of only one Pru. Our findings suggest that hRpn13 can form bidentate interactions with ubiquitinated substrates by binding to both ubiquitin chains and disordered sequences of substrates. Testing this model by solution nuclear magnetic resonance revealed Pru to bind weakly to various peptides, concurrent binding with ubiquitin, and ENT displacement by hRpn2, the latter required for substrate handoff to the proteasome ATPases.
人源Rpn13中一种类普列克底物蛋白的泛素受体（Pru）结构域可结合泛素和蛋白酶体亚基hRpn2。在此，我们报道了Pru与重组人源Rpn13极端N端（ENT）氨基酸结合的晶体结构。ENT呈U形，一侧为天然序列，其中M1埋于以Pru的W108为中心的口袋中，另一侧为非天然序列，其主链与晶格中相邻的Pru形成氢键。即使仅包含一个Pru，这些ENT:Pru相互作用在分子动力学模拟中也是稳定的。我们的研究结果表明，hRpn13可通过与泛素链和底物的无序序列结合，与泛素化底物形成双齿相互作用。通过溶液核磁共振对该模型进行测试，结果显示Pru与各种肽的结合较弱，与泛素同时结合，且hRpn2可置换ENT，后者是底物传递给蛋白酶体ATP酶所必需的。
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