军事医学2025，Vol.49Issue(8)：598-603,6.DOI:10.7644/j.issn.1674-9960.2025.08.006
有机磷水解酶突变体的SUMO融合表达及活性评价
SUMO-fusion expressions and catalytic efficiency of an organophosphate hydrolase mutant
刘壮 1翟亚楠 1王顺业 1马铭 1王梓洋 1刘艳芹 1高翔 1高静1
作者信息
- 1. 军事科学院军事医学研究院,北京 100850
- 折叠
摘要
Abstract
Objective To heterologously express and purify the small ubiquitin-like modifier(SUMO)tag fused organo-phosphorus hydrolase mutant H257Y/L303T(YT),namely SUMO-YT,and evaluate its hydrolytic capacity against ethyl paraoxon and soman.Methods The SUMO tag encoding gene was constructed at the N-terminus of the YT encoding gene with a linker sequence via enzyme digestion and ligation before SUMO-YT was expressed in Escherichia coli.SUMO-YT and YT were purified through ammonium sulfate precipitation and affinity chromatography to obtain high-purity target proteins.The activity and kinetic parameters of the recombinant enzymes were examined using ethyl paraoxon and soman as substrates.Results The system for expression and purification of recombinant enzymes was established,yielding SUMO-YT and YT,and the former exhibited more significantly enhanced hydrolytic efficiency than the latter,with catalytic rates 11-fold higher for paraoxon and 4.4-fold higher for soman.At 37 ℃ and pH 7.2,SUMO-YT reduced the inhibition rate of acetylcholinesterase(AChE)by soman from 100％to 45.7％within 3 minutes,whereas YT reduced it to no more than 80％.Conclusion The high-activity recombinant SUMO-YT is prepared.SUMO tag fusion can significantly enhance the hydrolytic capacity of YT against ethyl paraoxon and soman.关键词
有机磷水解酶/SUMO标签/乙酰胆碱酯酶/乙基对氧磷/梭曼/动力学参数Key words
organophosphate hydrolase/SUMO-Tag/acetylcholinesterase/ethyl paraoxon/soman/kinetic parameters分类
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刘壮,翟亚楠,王顺业,马铭,王梓洋,刘艳芹,高翔,高静..有机磷水解酶突变体的SUMO融合表达及活性评价[J].军事医学,2025,49(8):598-603,6.
