中国生物工程杂志

Objective To optimize the renaturation procedure of denatured LexA, prepare the repressor LexA from Pseudomonas aeruginosa (PA), which have the satisfactory biologic activity. Methods The LexA was renatured by the GSH/GSSG dilution method, and the renatured protein were purified by Ni2+ chelate affinity chromatography and gel filtration chromatography, following desalination by Sephadex G-25 gel column. The renaturation result were detected by the native polyacrylamide gel electrophoresis and RP-HPLC.The immunological activity of all LexA proteins, including the denatured , renatured protein and the renatured protein that was treated with the DTT, were determined by Western blot. Results The renatured LexA appears both monomer and multimer, which is confirmed by the native polyacrylamide gel electrophoresis analysis and RP-HPLC. Gel retardation experiments shows that the renatured LexA have satisfactory biologic activity.