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. 2016 Dec 5;18(1):138–149. doi: 10.15252/embr.201643206

Figure 4. MltE interacts with the TssM periplasmic domain.

Figure 4

  • A, B
    Bacterial two‐hybrid assay. BTH101 reporter cells producing the indicated proteins or domains fused to the T18 or T25 domain of the Bordetella adenylate cyclase were spotted on X‐gal indicator plates. The blue color of the colony reflects the interaction between the two proteins. TolB and Pal are two proteins known to interact but unrelated to the T6SS or the MltE proteins. The experiment was performed in triplicate and a representative result is shown.
  • C
    Co‐immunoprecipitation assay. The solubilized lysates from 2 × 1010 Escherichia coli K‐12 W3110 cells co‐expressing the indicated FLAG‐tagged TssMP variants (exported in the periplasm) and VSV‐G‐tagged MltE protein (Total, T) were subjected to immunoprecipitation on anti‐FLAG‐coupled agarose beads. The immunoprecipitated material (IP) was subjected to 12.5% acrylamide SDS–PAGE and immunodetected with anti‐FLAG (upper panel, TssM domains) and anti‐VSV‐G (lower panel, MltE) antibodies. Molecular weight markers (in kDa) are indicated on the left. The experiment was performed in triplicate and a representative result is shown.