Figure 2. Native-PAGE, SDS/PAGE and Western blot analyses of the 26S proteasome from T. bernacchii.

(A) Native-PAGE of the purified T. bernacchii 26S (26S b), immunoblotted against 19S subunit Rpt1 or 20S subunits β₅/β₁. The commercially available 26S from human erythrocytes (26S h) was used as positive control. (B) Native-PAGE of 26S b followed either by in-gel detection of CT-like activity (β₅ subunit), using the fluorogenic substrate LLVY, than Coomassie-blue stained. 26S h and 20S h were used as positive controls. The main proteasome species are indicated: 19S–20S–19S, doubly capped 26S; 19S–20S, singly capped 26S; 19S (regulatory complex) and 20S (catalytic core), free particles. (C) SDS/PAGE analysis of 26S b in comparison with 26S h and 20S h used as controls. (D) SDS/PAGE of 26S b immunoblotted against 20S subunits β₅/β₁. The 26S h was used as control. The results were representative of three independent experiments on three different protein preparations.