Probing requirements to bind a 42kDa cellular target and accumulate high molecular weight polyubiquitinated proteins.

Abstract

(A) Chemical structures of candidate inhibitors. (B) Precleared ES2 cell lysate was incubated with compounds (20 μM) or vehicle (DMSO, 1:100) for 45 min and then with RA190B (5 μM) for 45 min at 4°C. Samples were subjected to SDS-PAGE, transfer to PVDF membrane, and after probing with HRP-streptavidin, binding was detected using chemiluminescence. (C) ES2 cells were treated with compounds (1 μM, 4 hr), lysed and the samples probed with ubiquitin or actin-specific antibody by Western blot.</p