Department of Biochemistry, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway, New Jersey 08854, USA.
J Biol Chem. 2010 Mar 12;285(11):8330-9. doi: 10.1074/jbc.M109.076786. Epub 2010 Jan 8.
A major fraction of intracellular protein degradation is mediated by the proteasome. Successful degradation of these substrates requires ubiquitination and delivery to the proteasome followed by protein unfolding and disassembly of the multiubiquitin chain. Enzymes, such as Rpn11, dismantle multiubiquitin chains, and mutations can affect proteasome assembly and activity. We report that different rpn11 mutations can affect proteasome interaction with ubiquitinated proteins. Moreover, proteasomes are unstable in rpn11-1 and do not form productive interactions with multiubiquitinated proteins despite high levels in cell extracts. However, increased levels of ubiquitinated proteins were found associated with shuttle factors. In contrast to rpn11-1, proteasomes expressing a catalytically inactive mutant (rpn11(AXA)) were more stable and bound very high amounts of ubiquitinated substrates. Expression of the carboxyl-terminal domain of Rpn11 partially suppressed the growth and proteasome stability defects of rpn11-1. These results indicate that ubiquitinated substrates are preferentially delivered to intact proteasome.
细胞内蛋白质降解的主要部分是由蛋白酶体介导的。这些底物的成功降解需要泛素化和递送至蛋白酶体，然后进行蛋白质展开和多泛素链的解体。酶，如 Rpn11，可拆分手多泛素链，突变会影响蛋白酶体的组装和活性。我们报告说，不同的 rpn11 突变可以影响蛋白酶体与泛素化蛋白的相互作用。此外，rpn11-1 中的蛋白酶体不稳定，尽管细胞提取物中水平很高，但不能与多泛素化蛋白形成有效相互作用。然而，发现与穿梭因子相关的泛素化蛋白水平增加。与 rpn11-1 相反，表达无催化活性突变体 (rpn11(AXA)) 的蛋白酶体更稳定，结合了非常大量的泛素化底物。Rpn11 的羧基末端结构域的表达部分抑制了 rpn11-1 的生长和蛋白酶体稳定性缺陷。这些结果表明，泛素化底物优先递送至完整的蛋白酶体。
