摘要
亲和胶合成实验以双环氧化合物 1,4 丁二醇 2 缩水甘油醚 (1,4 butanedioldiglycidylether)为活化体及连接臂 ,在硼氢化钠 (NaBH)存在的碱性条件下 ,将载体SepharoseCL%D 4B与雷诺普利 (lisinopril)共价连接在一起 ,成功合成亲和层析胶 ,并利用亲和层析胶对猪肺血管紧张素转换酶 (angiotensinconvertingenzyme ,ACE)进行分离提纯 .猪肺组织匀浆经 1 6～ 2 6mol/L硫酸铵分级沉淀、透析平衡、亲和柱分离等步骤 ,从 2 0 0 g猪肺中提纯得到 0 79mgACE蛋白 ,酶活力回收 11 9% ,比活力 38 8U/mg .与层析前的酶液比较 ,亲和层析一步提纯可达 2 6 4倍 ;与肺匀浆液比较提纯达 80 8倍 .SDS 聚丙烯酰胺凝胶电泳可见 ,提纯的猪肺ACE为一条带 ,分子质量约为
A method for preparing biospecific affinity chromatography agarose for angiotensin converting enzyme is reported. Bisoxiranes, 1,4 butanediol diglycidyl ether, has been used for introducing reactive oxirane groups into Sepharose CL 4B, and coupling to ACE selective inhibitor——lisinopril in anion conditions including NaBH as deoxidizer. 0 79 mg ACE protein was purified from 200 g hog lung homogenate by 1 6～2 6 mol/L ammonium sulfate fractionation, dialysis, buffer balance and affinity chromatography. The recovery of ACE activity was 11 9% with specific activity of 38 8 U/mg protein and the enzyme was purified 808 fold in comparison with the homogenate supernatant of hog lungs, specially, only one step of affinity chromatography, the enzyme was purified 264 fold. Purified ACE showed a single band after migrated identically in sodium dodecyl sulphate polyacrylamide gel electrophoresis and given an apparent molecular mass about 180 ku.
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
2000年第5期544-547,共4页
Progress In Biochemistry and Biophysics
基金
国家新药研究基金!(96 90 1 0 5 85 ) .
关键词
分离
纯化
血管紧张素转换酶
亲和层析法
affinity agarose preparation, purification, angiotensin converting enzyme
