Figure 1. GlpG and Rhom7 are active rhomboids.

1. Topology of GlpG, Rhom7 and the artificial substrate with a maltose‐binding protein (MBP) domain, triple‐FLAG tag (3xFLAG), the TMD of P. stuartii TatA and a thioredoxin domain (Trx).
2. Western blot analysis (probing with an anti‐FLAG mAb) to detect cleavage of the artificial substrate by wild‐type (WT)/inactive (S²⁰¹A) GlpG encoded on pBAD33 with/without arabinose (Ara.).
3. Western blot analysis to detect cleavage of the artificial substrate by wild‐type (WT)/modified (S¹³³A or H¹⁸⁷A) Rhom7 encoded on pBAD33 with/without arabinose (Ara.).
4. Activity of Rhom7 with/without its 7^th TMD and/or C‐terminal domain.

Data information: In (B‐D), rhomboid substrates that are uncleaved, cleaved by GlpG or cleaved by Rhom7 are marked by black, red and blue arrows, respectively. Controls, empty pBAD33 (empty) and wild‐type Shigella sonnei (Ss). RecA, loading control. Source data are available online for this figure.