血管紧张素转换酶纯化与性质研究

Purification and Characterization of Angiotensin Converting Enzyme

为了深入了解猪肺血管紧张素转换酶 (angiotensin converting enzyme,ACE)的性质和功能 ,对猪肺 ACE的分离纯化以及部分酶学性质进行了研究 .猪肺组织匀浆经 1 .6～ 2 .6mol/L硫酸铵分级沉淀等步骤后 ,利用亲和胶进行亲和层析分离 .2 0 0 g猪肺组织中提纯出 0 .79mg ACE,比活力 38.8U/mg,SDS- PAGE电泳鉴定为一条带 ,分子量约 1 80 k D,等电点 (p I)为 p H4.5,糖含量约 2 3.6% ,氨基酸组成分析发现猪肺 ACE分子中含有 1 346个氨基酸 ,其中酸性氨基酸含量较高 ,碘乙酸的修饰结果表明猪肺 ACE中巯基基团未参与酶的催化反应 .酶反应动力学结果显示 ,ACE催化 Fa PGG底物反应时的最适 p H大约为 p H 7.6,反应活化能 Ea=4.37× 1 0 4 J/mol,酶活性部位附近的组氨酸和具有类似 α-氨基性质的氨基酸可能参与了 ACE催化反应 .有关猪肺 ACE的基本生化性质、氨基酸组成以及酶学性质的结果 ,为今后深入研究奠定了基础 .

In order to get a full comprehension on properties of hog angiotensin converting enzyme(ACE),the research on the purification and characterization of hog ACE was carried out.After hog lung homogenate processed with 1.6-2.6 mol/L ammonium sulfate fractionation and so on,0.79 mg angiotensin converting enzyme(ACE) protein was purified from 200 g hog lung tissue using affinity chromatography columm.The purified ACE specific activity reached 38.8 units/mg,sodium dodecyl sulphate polyacrylamide gel electrophoresis(SDS PAGE)detecting showed a single band and gave an apparent M r about 180 kD.The isoelectric point(p I )of ACE was about pH4.5 and saccharide content was about 23.6%.1 346 amino acid residues were found to contain in an ACE molecule concluding high level of acid amino acid residues through amino acid composition analysis.The optimum pH of ACE was bout 7.6 and the histidine and α amino amino acid analogs were supposed to partake in the enzyme catalytic reaction by enzyme reaction kinetics experiments,and calculated that the reaction activation energy of ACE catalyzing FaPGG was 4.37 ×10 4 J/mol.The effects of amino acid composition,characterization and enzymic property of ACE established a foundation for further research.