摘要
目的 提供一个简便的甲状腺过氧化物酶 (TPO)的纯化方法 ,并研究其抑制物对酶活性的影响。方法 用差速离心、胰蛋白酶与去氧胆酸钠增溶、硫酸铵盐析及 DE5 2、Sephadex G10 0和 DEAE- Sephadex A5 0三步柱层析法从猪甲状腺中提纯 TPO。比较不同浓度的氟化钠 (Na F)、硫氰化钾 (KSCN )、丙基硫脲嘧啶 (PTU )及叠氮钠 (Na N3 )对 TPO活性的影响。结果 TPO提纯终产物的纯化倍数为 2 98.6 ,其 RZ值为 0 .2 7。 Na N3 、PTU、KSCN和 Na F抑制 TPO活性 ,并且随着其浓度的升高 ,抑制作用增强 ,抑制常数分别为 4.4,3.2 ,2 .6和1.8。结论 提供的 TPO纯化方法简便、可靠。 Na N3 、PTU、KSCN和 Na F对
Objective To purify thyroid peroxidase (TPO) from porcine thyroid tissue by an improved and simplified method and to study the effects of the inhibitors on activity of the enzyme.Methods The membrane- bound enzyme was solubilized by trypsin plus detergent. After precipitation with ammonium sulfate, the enzyme was purified by a series of column treatments, including ion-exchange chromatography on DEAE-cellulose, gel filtration through Sephadex G100 and DEAE-Sephadex A50 chromatography. We also compared the effects on activity of TPO with various concentrations of thiocyanide(KSCN), fluoride(NaF), azide(NaN 3) and propylthiouracil(PTU).Results The final product of purification had a value for RZ of 0.27 and for purified fold of 298.6. TPO was inhibitied by NaN 3, PTU, KSCN and NaF.With various inhibitory concentrations sigmoidal inhibition curves were obtained. The inhibition constants, obtained from the curves,were 4.4(NaN 3), 3.2(PTU), 2.6(KSCN) and 1.8(NaF).Conclusions The results suggest that the method of purification is improved and depended.The effects of the inhibitors on activity of TPO are lowered in order(NaN 3>PTU>KSCN>NaF).
出处
《中国地方病学杂志》
CAS
CSCD
北大核心
2001年第5期328-331,共4页
Chinese Jouranl of Endemiology
基金
辽宁省教委基金资助项目 ( 96 1131115 2 )
