Overproduction, purification and characterization of GroES and GroEL from thermophilic Bacillus stearothermophilus.

To facilitate purification of the two chaperonins GroES and GroEL encoded by the thermophilic Bacillus stearothermophilus, an Escherichia coli strain was constructed in which the geoESL operon was replaced by that of B. stearothermophilus. This strain is perfectly viable, demonstrating that the B. stearothermophilus operon is functionally interchangeable with that of E. coli. To increase the amount of GroES, the groES gene was fused to an IPTG-inducible promoter. Both proteins GroES and GroEL, were purified from E. coli using the standard protocol with some modifications. This method should be applicable in all cases where a foreign groE operon can substitute that of E. coli. A preliminary characterization of GroEL, revealed that it has the same secondary structural elements as the E. coli homologue, but its thermodynamic stability is significantly increased.