Effects of chaperone co-expression on heterologous solubility expression of thermophilic cyclodextrin glucosetransferase

Abstract:

[Objective] To improve the soluble expression level and enzyme activity of CGTase in prokaryotic expression system, the expression conditions were optimized. [Methods] The cgt gene was cloned from Geobacillus sp. B1 and cloned into expression vector pET-28a(+). The optimum induction temperature was selected by enzyme activity assay and SDS-PAGE. The molecular chaperone co-expression system was constructed and the optimum molecular chaperone vector was screened. [Results] The results was described as follows: the cgt gene was amplified and cloned into vector pET-28a(+) successfully, the enzyme activity and soluble expression level of CGTase was highest when induced at 25 °C; in the molecular chaperone co-expression system, the five vectors containing different molecular chaperones (pKJE8, pKJE7, pGro7, pTf16 and pG-Tf2) improved and the enzyme activity and soluble expression level of CGTase in varying degrees when co-expression with recombinant plasmid pET-28a(+)-ompA-cgt, and pKJE8 was confimed to contain the optimum molecular chaperones combination and the enzyme activity of CGTase improved 48.6%. When L-Arabinose concentration of 0.5 g/L, molecular chaperone plasmid pKJE8 made extracellular enzyme activity increased 68.5%. [Conclusion] These results can provide a potential value for further studies of CGTase.