FIG 4.

Overproduction of Ph2119 endolysin in E. coli Tuner(DE3) cells. Shown is SDS-PAGE (12.5%) of the cellular proteins from 100-μl cultures of bacteria. (A) E. coli Tuner(DE3)(pMP20) before induction (lane 1) and 4 h after induction (1 mM IPTG) (lane 2). The position of Ph2119 endolysin is indicated by an arrow. (B) Successive steps in the purification of Ph2119 endolysin on a column with Ni-NTA agarose. Lanes: 1, total protein; 2, soluble protein after heat treatment (15 min at 75°C); 3, flowthrough; 4, 1st wash with 10 mM imidazole in buffer NPI; 5, 2nd wash with 20 mM imidazole in buffer NPI; 6, Ph2119 endolysin eluted with 250 mM imidazole in buffer NPI; M, molecular masses of reference proteins (Thermo Scientific) (in kilodaltons).