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. 2022 Jan 21;20(1):e3001523. doi: 10.1371/journal.pbio.3001523

Fig 3. TolC is a substrate of TamA.

Fig 3

(A) Structural map as in Fig 1A. The boundaries of the transmembrane domain are indicated below the structural map, and the positions of each cysteine substitution are indicated above. (B) Ribbon diagram of the TolC β-barrel domain (PDB: 1EK9). Each cysteine substitution is highlighted in yellow. Transmembrane β-strands are coloured as in panel (A). (C-D) Membranes were isolated from ΔtamA cells expressing the indicated tolC and tamA variants. Peripheral proteins (DnaK) were separated from integral proteins (BamA) by Triton X-114 extraction. Total membranes (T), aqueous phases (A), and detergent-enriched phases (D) were analysed by immunoblotting after nonreducing SDS-PAGE (TamA) or reducing SDS-PAGE (BamA and DnaK). White triangles indicate TamA–TamA disulphide interactions and black triangles indicate TamA–TolC interactions. This experiment was performed in biological triplicate (from 3 independent membrane preparations). Uncropped images are presented in S1 Raw Images; original immunoblots (including relevant replicates) are presented in S3 Raw Images.