Effects of different signal peptides and chemical penetrators on extracellular production of recombinant cyclodextrin glycosyltransferase
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    Abstract:

    [Objective] Effects of signal peptides and chemical penetrators on extracellular secretion of cyclodextrin glycosytransferase (CGTase) in recombinant Escherichia coli strain BL21(DE3) was observed in this study in order to lay foundation for fermentation technology of CGTase. [Methods] The CGTase gene was cloned by PCR with genomic DNA of Geobacillus sp. CHB1 as template. Four recombinant plasmids were constructed which included the signal peptide of Geobacillus sp. CHB1, OmpA, PelB and no signal peptide. The recombinant plasmids were respectively transformed into E. coli BL21(DE3) and then induced to express. The extracellular enzyme activities of the target proteins were analyzed and measured to screen the best signal peptide. Chemical penetrators like glycine, SDS, Triton X-100 and Tween 80 were added into the culture to determine the effect of chemical additives on extracellular secretion of recombinant CGTase. [Results] The results showed that these four recombinant plasmids could successfully express CGTase in E. coli BL21(DE3) and OmpA was the optimal signal peptide. Extracellular enzyme activity induced by OmpA could reach 7.44 U/mL which were 2.04- and 11.27-folds of PelB and CHB1, respectively. No extracellular CGTase activity was detected in E. coli BL21(DE3) with the no signal peptide. Extracellular enzyme activities induced by OmpA were 9.27 U/mL and 9.75 U/mL by addition of 0.6% glycine and 0.3% Triton X-100, respectively, and it could reach 14.27 U/mL by synergistic action of glycine and Triton X-100 after 48 h. However, SDS and Tween 80 had obvious inhibition on extracellular CGTase activity. [Conclusion] The extracellular CGTase activity in E. coli BL21(DE3) induced by OmpA could reach the highest by addition of 0.6% glycine and 0.3% Triton X-100.

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CAI Hai-Song, LIN Xiao-Xu, GUO Yong-Hua, CHEN Long-Jun, ZHANG Hui, QIU Hong-Duan, LIN Xin-Jian, CHEN Ji-Chen. Effects of different signal peptides and chemical penetrators on extracellular production of recombinant cyclodextrin glycosyltransferase[J]. Microbiology China, 2017, 44(3): 601-610

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  • Online: March 09,2017
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