摘要
人脑乙酰胆碱酯酶的全长cDNA序列克隆到真核高效表达载体pcDNA3.1中 ,并将pcDNA AChE转染人胚肾细胞株 2 93细胞 ,进行rhAChE的暂时表达 .真核细胞表达的rhAChE的生化性质与天然人脑AChE十分相似 .rhAChE的Km 值约为 137μmol L ;有过量底物抑制现象 ;可被胆碱酯酶抑制剂huperzineA和eserine抑制 (IC50 分别为 2 5× 10 -8mol L和 1 0× 10 -7mol L) ;肟类化合物HI 6 (10 -4 mol L)可以有效地重活化被sarin(10 -6mol L及 10 -7mol L)抑制的rhAChE ,4h内重活化率分别达 86 %和 97% .rhAChE反复冻融 3次 ,酶活性没有损失 .
The encoding sequence of human brain acetylcholinesterase was subcloned into an eukaryotic expression vector pcDNA 3.1 and then transfected into human embryonic kidney cell line 293 for expression of recombinant human acetylcholinesterase. The biotoxicological properties of recombinant acetylcholinesterase and native acetylcholinesterase were very much alike. The K m value of rhAChE was 137 μmol/L; rhAChE exhibited the excess substrate inhibition. Inhibitor huperzine A and eserine could inhibit rhAChE with IC 50 of 2 5×10 -8 mol/L and 1×10 -7 mol/L respectively. HI 6(10 -4 mol/L) reactivated the sarin(10 -6 mol/L and 10 -7 mol/L) inhibited rhAChE with reactivation rates of 86% and 97% .The rhAChE could stand repeated freezing and thawing for three times without loss of enzyme activity.
出处
《中国生物化学与分子生物学报》
CAS
CSCD
北大核心
2002年第1期44-49,共6页
Chinese Journal of Biochemistry and Molecular Biology
基金
全军"九五"医学科研基金重点项目 ( 96 Z 0 16 )