Department of Chemistry, The Scripps Research Institute, 130 Scripps Way, Jupiter, FL 33458, USA.
Department of Molecular Medicine, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Cell Chem Biol. 2020 Nov 19;27(11):1371-1382.e6. doi: 10.1016/j.chembiol.2020.08.007. Epub 2020 Aug 27.
Rpn13 is one of several ubiquitin receptors in the 26S proteasome. Cys88 of Rpn13 has been proposed to be the principal target of RA190, an electrophilic small molecule with interesting anti-cancer activities. Here, we examine the claim that RA190 mediates its cytotoxic effects through engagement with Rpn13. We find no evidence that this is the case. In vitro, RA190 is has no measurable effect on any of the known interactions of Rpn13. In cellulo, we see no physical engagement of Rpn13 by RA190, either on C88 or any other residue. However, chemical proteomics experiments in two different cell lines reveal that dozens of other proteins are heavily engaged by RA190. Finally, increasing or reducing the level of Rpn13 in HeLa and melanoma cells had no effect on the sensitivity of HeLa or melanoma cells to RA190. We conclude that Rpn13 is not the physiologically relevant target of RA190.
Rpn13 是 26S 蛋白酶体中几种泛素受体之一。Rpn13 的 Cys88 被提议为 RA190 的主要靶标，RA190 是一种具有有趣抗癌活性的亲电小分子。在这里，我们研究了 RA190 通过与 Rpn13 结合来介导其细胞毒性作用的说法。我们没有发现事实如此的证据。在体外，RA190 对 Rpn13 的任何已知相互作用都没有可衡量的影响。在细胞内，我们没有看到 RA190 与 Rpn13 的任何其他残基在 C88 上发生物理结合。然而，在两种不同的细胞系中的化学蛋白质组学实验表明，数十种其他蛋白质与 RA190 大量结合。最后，增加或减少 HeLa 和黑色素瘤细胞中 Rpn13 的水平对 HeLa 或黑色素瘤细胞对 RA190 的敏感性没有影响。我们得出结论，Rpn13 不是 RA190 的生理相关靶标。
