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. Author manuscript; available in PMC: 2018 Sep 12.
Published in final edited form as: Nat Microbiol. 2018 Mar 12;3(4):481–493. doi: 10.1038/s41564-018-0122-x

Figure 6. The TRiC chaperonin folds σ3 into a native, assembly-competent conformation.

Figure 6

a, Schematic of σ3 folding and assembly experiments. b, SDS-PAGE of 35S-met-labeled σ3 immunoprecipitated from RRLs using a conformation-specific antibody (10C1 monoclonal antibody) or isotype control. Where indicated, RRLs were TRiC-depleted and reconstituted with purified hTRiC (+, 0.25 μM) (10% input loaded into lanes 1,4,7). c, SDS-PAGE of 35S-met-labeled σ3 translated in RRLs mock-depleted, TRiC-depleted, or TRiC-depleted and reconstituted with purified hTRiC (+, 0.25 μM) and incubated with proteinase K (2.5 μg/mL final concentration) for the times shown. d, SDS-PAGE and colloidal blue stain of T1L reovirus virions and ISVPs. e, SDS-PAGE of ISVPs recoated with 35S-met-labeled σ3 translated in RRLs and immunoprecipitated using a μ1δ-specific antibody, which is specific to the μ1 species on ISVPs. Where indicated, RRLs were TRiC-depleted and reconstituted with purified hTRiC (+, 0.125 μM; ++, 0.50 μM) (20% input loaded into lanes 1,4,7,10). Arrows in SDS-PAGE gels indicate full-length σ3. In b–e, three independent experiments were conducted with similar results. f, Model of the TRiC-σ3 folding and assembly pathway.