Kang Yang, Zhang Naixia, Koepp Deanna M, Walters Kylie J
Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA.
J Mol Biol. 2007 Jan 26;365(4):1093-101. doi: 10.1016/j.jmb.2006.10.056. Epub 2006 Oct 21.
Ubiquitin receptor proteins play an important role in delivering ubiquitylated protein substrates to the proteasome for degradation. HHR23a and hPLIC2 are two such ubiquitin receptors that contain ubiquitin-like (UBL) domains, which interact with the proteasome, and ubiquitin-associated (UBA) domains, which interact with ubiquitin. Depending on their abundance UBL/UBA family members can either promote or inhibit the degradation of other proteins, which suggests their participation in the delivery of substrates to the proteasome is highly regulated. In previous work, we determined UBL/UBA domain interactions to promote intramolecular interactions in hHR23a that are abrogated with the addition of either ubiquitin or the proteasome component S5a. In yeast, we determined the hHR23a ortholog (Rad23) to interact with another UBL/UBA family member (Ddi1) and to bind a common tetraubiquitin chain. Here, we use NMR spectroscopy to reveal that hHR23a interacts with hPLIC2 via UBL/UBA domain interactions and to map their binding surfaces. In addition, we demonstrate that these two proteins associate in mammalian cells. Intriguingly, inhibition of the proteasome mitigates hHR23a/hPLIC2 interaction.
泛素受体蛋白在将泛素化的蛋白质底物递送至蛋白酶体进行降解过程中发挥着重要作用。HHR23a和hPLIC2就是这样两种泛素受体，它们含有与蛋白酶体相互作用的类泛素（UBL）结构域以及与泛素相互作用的泛素相关（UBA）结构域。根据其丰度，UBL/UBA家族成员既可以促进也可以抑制其他蛋白质的降解，这表明它们参与底物向蛋白酶体的递送受到高度调控。在之前的工作中，我们确定了UBL/UBA结构域相互作用可促进hHR23a中的分子内相互作用，而添加泛素或蛋白酶体组分S5a可消除这种相互作用。在酵母中，我们确定hHR23a的直系同源物（Rad23）与另一个UBL/UBA家族成员（Ddi1）相互作用并结合一条常见的四聚泛素链。在此，我们利用核磁共振光谱揭示hHR23a通过UBL/UBA结构域相互作用与hPLIC2相互作用，并绘制它们的结合表面。此外，我们证明这两种蛋白质在哺乳动物细胞中相互关联。有趣的是，蛋白酶体的抑制减轻了hHR23a/hPLIC2的相互作用。
