Identification of the minimal AT module permitting secretion of heterologous proteins to the culture supernatant fraction. (A) Schematic of ESAT6-Pet-BP protein fusion and some truncations created to determine the minimal C-terminal Pet fragment capable of ESAT-6 secretion. The Δ*1, Δ*2, Δ*6, Δ*17 and Δ*20 Pet truncations are shown while for simplicity the intermediate variants Δ*3–Δ*5, Δ*7–Δ*16, Δ*18 and Δ*19 are omitted. Abbreviations are the same as in Figure 1. (B and C) Detection by western immunoblotting of ESAT6-Pet (B) and ESAT6-Pic (C) chimeras expressed in E. coli TOP10. The TCA-precipitated culture supernatants were analysed by SDS-PAGE and probed with polyclonal anti-ESAT6. The equivalent OM fractions demonstrating the presence of the cleaved β-barrel in the OM are shown in Additional file 6: Figure S5.