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On-line Access: 2024-08-27
Received: 2023-10-17
Revision Accepted: 2024-05-08
Crosschecked: 2021-01-29
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Citations: Bibtex RefMan EndNote GB/T7714
Liujia SHI, Fangfang YANG, Yanyan XU, Shoufeng WANG. Expression of Drosophila melanogaster acetylcholinesterase (DmAChE) gene splice variants in Pichia pastoris and evaluation of its sensitivity to organophosphorus pesticides[J]. Journal of Zhejiang University Science B, 2021, 22(3): 204-213.
@article{title="Expression of Drosophila melanogaster acetylcholinesterase (DmAChE) gene splice variants in Pichia pastoris and evaluation of its sensitivity to organophosphorus pesticides",
author="Liujia SHI, Fangfang YANG, Yanyan XU, Shoufeng WANG",
journal="Journal of Zhejiang University Science B",
volume="22",
number="3",
pages="204-213",
year="2021",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B2000525"
}
%0 Journal Article
%T Expression of Drosophila melanogaster acetylcholinesterase (DmAChE) gene splice variants in Pichia pastoris and evaluation of its sensitivity to organophosphorus pesticides
%A Liujia SHI
%A Fangfang YANG
%A Yanyan XU
%A Shoufeng WANG
%J Journal of Zhejiang University SCIENCE B
%V 22
%N 3
%P 204-213
%@ 1673-1581
%D 2021
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B2000525
TY - JOUR
T1 - Expression of Drosophila melanogaster acetylcholinesterase (DmAChE) gene splice variants in Pichia pastoris and evaluation of its sensitivity to organophosphorus pesticides
A1 - Liujia SHI
A1 - Fangfang YANG
A1 - Yanyan XU
A1 - Shoufeng WANG
J0 - Journal of Zhejiang University Science B
VL - 22
IS - 3
SP - 204
EP - 213
%@ 1673-1581
Y1 - 2021
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B2000525
Abstract: acetylcholinesterase (AChE) is a key enzyme used to detect organophosphorus pesticide residues by the enzyme inhibition method. An accidental discovery of a mutant strain with AChE activity was made in our laboratory during the process of AChE expression by Pichia pastoris. The pPIC9K-Drosophila melanogaster acetylcholinesterase (DmAChE)-like expression vector was constructed by codon optimization of this mutant strain, which was transformed into P. pastoris GS115, and positive clones were selected on yeast peptone dextrose (YPD) plate with G418 at 4.0 mg/mL. The GS115-pPIC9K-DmAChE-like strain was subjected to 0.5% methanol induction expression for 120 h, with a protein band at 4.3 kDa found by the tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) pattern of the fermentation supernatant. After preliminary purification by ammonium sulfate precipitation, the enzyme activity was detected to be 76.9 U/(mL⋅min). In addition, the pesticide sensitivity test proved that DmAChE-like is selective and sensitive to organophosphorus pesticides.
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