Stable expression and rapid purification of Escherichia coli GroEL and GroES chaperonins.

An Escherichia coli expression vector pRE (P. Reddy, A. Peterkofsky, and K. McKenney, 1989, Nucleic Acids Res. 17, 10473-10488), originally developed for the cloning and expression of lethal genes, was used for cloning and hyperexpression of GroEL and GroES genes. Regulated gene expression is achieved in the pRE vector under the tight control of the lambda PL promoter. Upon induction of the promoter, stable expression of GroEL to about 60% of the total cell protein was observed. Similarly, stable expression of GroES to about 40% of the total cell protein was achieved. GroES was found to be a heat-stable protein while GroEL was not. Both GroE chaperonins were purified in a single chromatographic step with a yield of about 100 mg GroEL and 25 mg GroES per liter of E. coli culture. GroE chaperonins purified by the protocols described here were active in the renaturation of urea-denatured rhodanese.