Figure 4
Expression of C. pseudotuberculosis Hsp60 protein and complementation assays. a) Expression of C. pseudotuberculosis Hsp60 protein in E. coli groEL44 mutant transformed with the recombinant vector pProEx-Hta:hsp60 (groEL44[pProEx-Hta/hsp60]); E. coli wild-type and mutant transformed with the empty vector (B178[pProEX-Hta] and groEL44[pProEX-Hta], respectively) were used as negative controls. Expression of recombinant protein was analyzed at 0, 20, 40 and 60 min after induction with IPTG. The rectangle indicates the protein expressed; b) Complementation assay after thermal stress. The growth of E. coli wild-type (B178[pProEX-Hta]), groEL44 mutant (groEL44[pProEX-Hta] and groEL44[pProEx-Hta/hsp60]) were evaluated. Cultures were serially diluted (10-1 to 10-6), spotted onto the surface of LB agar plates (with and without 1 mM IPTG), and incubated at 30 or 42°C for 18 h; c) Complementation analysis of growth of the γc Ib2 phage in E. coli groEL44 mutant. The growth of the phage was evaluated in E. coli wild-type (B178[pProEX-Hta]) and groEL44 mutant (groEL44[pProEX-Hta] and groEL44[pProEx-Hta/hsp60]). The cultures induced with IPTG, plated at the dilutions from10-1 to 10-4, were incubated at 30°C for 18 h.