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Figure 1 | BMC Microbiology

Figure 1

From: Role of EscU auto-cleavage in promoting type III effector translocation into host cells by enteropathogenic Escherichia coli

Figure 1

Efficient translocon and effector secretion is dependent on EscU auto-cleavage. (A): Immunoblot demonstrating EscU variant cleavage status within whole cell lysates. The blots were imaged separately to get representative signals for the auto-cleavage products. A longer exposure was used for the 39 kDa protein species. (B) Left: trans-complementation of ΔescU with pJLT21 restores EspA (~20 kDa), EspB and EspD (co-migrate ~30 kDa) secretion to wild type levels (the identity of these dominant protein species has been previously determined using protein sequencing [36], and here by immunoblotting [right panel]). EspC is an abundant type 5 secreted protein. Bovine serum albumin (BSA) was added to collected secreted protein fractions as a carrier protein to assist in the precipitation of proteins. A molecular weight standard is in the left most lane. Right: immunoblot analyses of secreted protein and whole cell lysate fractions from bacterial strains used in panel A (as indicated). The respective secreted protein fractions were diluted 20 fold prior to SDS-PAGE. (C) Left: secreted protein fractions derived from ΔescNΔescU double mutant strains with the indicated plasmids. Right: Immunoblot analysis of secreted protein fractions. DnaK, an abundant non-secreted cytoplasmic protein, was used as a gel loading control (when needed) or to assess cytoplasmic contamination of secreted fractions or non-specific bacterial lysis. All samples were diluted 20 fold as in panel B. All experiments within the panels were performed twice and representative images are shown.

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