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Surface display and bioactivity of Bombyx mori acetylcholinesterase on Pichia pastoris

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A Pichia pastoris (P. pastoris) cell surface display system of Bombyx mori acetylcholinesterase (BmAChE) was constructed and its bioactivity was studied. The modified Bombyx mori acetylcholinesterase gene (bmace) was fused with the anchor protein (AGa1) from Saccharomyces cerevisiae and transformed into P. pastoris strain GS115. The recombinant strain harboring the fusion gene bmace-AGa1 was induced to display BmAChE on the P. pastoris cell surface. Fluorescence microscopy and flow cytometry assays revealed that the BmAChE was successfully displayed on the cell surface of P. pastoris GS115. The enzyme activity of the displayed BmAChE was detected by the Ellman method at 787.7 U/g (wet cell weight). In addition, bioactivity of the displayed BmAChE was verified by inhibition tests conducted with eserine, and with carbamate and organophosphorus pesticides. The displayed BmAChE had an IC50 of 4.1761028 M and was highly sensitive to eserine and five carbamate pesticides, as well as seven organophosphorus pesticides. Results suggest that the displayed BmAChE had good bioactivity.
Jie-Xian Dong , Xi Xie , Young-Sheng He , Ross C. Beier , Yuan-Ming Sun , Zhen-Lin Xu , Wei-Jian Wu , Yu-Dong Shen , Zhi-Li Xiao , Li-Na Lai , Hong Wang , Jin-Yi Yang
Bombyx mori , Pichia pastoris , Saccharomyces cerevisiae , acetylcholinesterase , carbamate pesticides , enzyme activity , flow cytometry , fluorescence microscopy , gene expression , gene fusion , genes , genetic transformation , inhibitory concentration 50 , organophosphorus pesticides
Plos One 2013 8 5 v.8 no.8
Journal Articles, USDA Authors, Peer-Reviewed
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