FIG. 2.

Nontoxic secretion of 6AC3HlyA hybrid to the extracellular medium of E. coli cultures. (A) Denaturing SDS-polyacrylamide gel (12%) stained with silver (1) showing the proteins secreted to the extracellular medium after 5 h of induction with 0.25 mM IPTG of E. coli HB2151(pVDL9.3) cells grown at 37°C and harboring pEHlyA (lane 1) or p6AC3HlyA (lane 2). Protein samples were prepared by adding to 50-μl portions of the culture supernatants an identical volume of 2× SDS-PAGE denaturing sample buffer (2). The mixtures were boiled for 5 min, and 10 μl of each sample was loaded per lane. For the molecular mass standards (M; Bio-Rad), 0.1 μg of each protein was loaded. (B) Growth curves in LB medium at 37°C of E. coli HB2151 cells harboring the indicated plasmids. The cultures were induced by 0.25 mM IPTG when the OD₆₀₀ was ∼0.4 (arrow).