CAJ | 학술논문

目的　重组人尿激酶原在大肠杆菌中过量表达时形成不溶物包涵体，需经体外变复性后才能获得生物活性。本文旨在提高包涵体中变性尿激酶原的复性效率。方法　通过对pH、温度、变性剂种类及浓度、蛋白浓度、以及巯基氧化还原对比率等的定性定量分析，研究重组人尿激酶原体外变复性的基本条件，并比较了添加一些非特异有效成分、脉冲稀释、梯度透析等方法对提高重组人尿激酶原体外变复性效率的作用。结果　确定了重组人尿激酶原体外变复性的基本方法，其复性效率可达30%左右。结论　不同的包涵体蛋白的体外变复性效率因蛋白的分子大小、二巯键数目、疏水程度等而异，对特定蛋白复性条件的优化可提高其复性效率。
목적　중조인뇨격매원재대장간균중과량표체시형성불용물포함체，수경체외변복성후재능획득생물활성。본문지재제고포함체중변성뇨격매원적복성효솔。 방법　통과대pH、온도、변성제충류급농도、단백농도、이급구기양화환원대비솔등적정성정량분석，연구중조인뇨격매원체외변복성적기본조건，병비교료첨가일사비특이유효성분、맥충희석、제도투석등방법대제고중조인뇨격매원체외변복성효솔적작용。 결과　학정료중조인뇨격매원체외변복성적기본방법，기복성효솔가체30%좌우。 결론　불동적포함체단백적체외변복성효솔인단백적분자대소、이구건수목、소수정도등이이，대특정단백복성조건적우화가제고기복성효솔。
Objective　Recombinant human pro-urokinase forms insoluble inclusion body when overexpressed in Escherichia coli. It must be denatured and renatured in vitro so that it can acquire activity. This study aimed at increasing the renaturation yield of denaturant pro-urokinase. Methods　We evaluated the basic renaturation conditions of pro-urokinase through qualitative and quantitative analysis of pH, temperature, denatured concentration, protein concentration, and the ratio of reduced and oxidized thiol reagents. We also compared the effects of nonspecific additives, step-wise dilution and urea gradient dialysis. Results　We defined the optimal conditions of pro-urokinase renaturation with a yield of about 20%-30%. Conclusion　Different recombinant denatured proteins have different renaturation conditions due to their different molecular sizes, molecular constructions, disulfide bond numbers, and hydrophobicity. The renaturation yield can be increased by optimizing the renaturation conditions of a specific protein.