The B. cereus toxins contain Sec-type signal peptides. (A) Sec-type signal peptide sequences of the Hbl, Nhe and CytK cytotoxin proteins from B. cereus ATCC 14579 predicted using SignalP. The predicted cleavage sites are marked with arrows and the hydrophobic regions are underlined. (B) Site-directed mutations introduced into the hydrophobic core of the signal peptide of Hbl B in this study. Western immunoblot analysis of Hbl B in culture supernatants and cell lysates of (C) B. cereus (Bc) NVH0075/95 and (D) the non-flagellated B. thuringiensis 407 flhA mutant (Bt407ΔflhA) transformed with pHT304-pXyl expressing native Hbl B and Hbl B with a mutated signal peptide sequence (Hbl Bmut). Negative controls are strains harbouring pHT304-pXyl empty vector (ctrl). Expression was induced by 20 mM xylose. Lane M: molecular weight marker.