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. 2021 Oct 20;12:739953. doi: 10.3389/fimmu.2021.739953

Figure 5.

Figure 5

Eri suppresses the ATPase activity of NLRP3. (A) HEK293T cells were transfected with indicated plasmids for 36 h. Immunoblot analysis of binding complexes isolated from BMDMs extracts incubated with Bio-Eri. (B) BMDMs were treated with LPS (1 µg/ml for 6 h) or Eri (5 nM for 6 h). Cells extracts were analyzed using SDD-AGE (upper panel) and SDS-PAGE (lower panel). (C) ATPase activity assay for purified NLRP3 in the presence of different dose of Eri. (D) ATPase activity assay for purified NLRP3, NLRC4, NLRP1 and NOD2 with or without presence of Eri (5 nM). (E) HEK293T cells were transfected with Flag-NLRP3 (left panel) or NLRP3 constructs with Walker A or Walker B motif mutation (right panel) for 36 h. Immunoblot analysis of binding complexes isolated from cells extracts incubated with Bio-Eri or free Eri. (F) ATP-binding assay for purified Flag-NLRP3 in the presence of Eri (5 nM). (G) ATPase activity assay for purified Flag-NLRP3 or mutants with or without presence of Eri (5 nM). All experiments were repeated at least three times. Bar graphs present means ± SD (**P < 0.01; n.s., not significant).