Soluble Expression and Purification of Q۵۹L Mutant L-asparaginase in the Presence of Chaperones in SHuffle™ T۷ strain

Background and Aims: Q۵۹L mutant of L-asparaginase enzyme from Escherichia coli (E. coli) has been introduced with lower side effects. This version of the enzyme might have potential applications in the treatment of leukemia patients. We utilized SHuffle T۷ strain of E. coli, to produce the mutant enzyme in the presence of chaperone molecules. Materials and Methods: Q۵۹LAsp gene was cloned into pET۲۸a expression vector, and two strains of E. coli (BL۲۱ DE۳ and SHuffle T۷ strains) were used to produce recombinant protein. In parallel, PG-Tf۲ plasmid was cloned into the same strains, and the effect of trigger factor chaperone and groELS chaperonines was studied. The his-tagged recombinant protein was expressed and purified using nickel affinity chromatography. The amount of recombinant protein which is produced in each condition was determined and compared. Results: The amount of soluble recombinant protein was enhanced in the presence of chaperones in both strains of E. coli. SHuffle T۷ strain produced more soluble recombinant protein in the soluble state than BL۲۱ DE۳ strain. So the best condition for the production of soluble recombinant Q۵۹L mutant protein was the use of PG-Tf۲ plasmid in the SHuffle T۷ strain. Conclusion: Application of the new strain SHuffle T۷, with chaperones simultaneously showed better results in the production of Q۵۹L mutant version of L-Asparaginase.

کلیدواژه ها:

Asparaginase ، Chaperone ، E. coli ، Shuffle ، آسپارژیناز ، لوسمی ، چاپرون ، سویه shuffle