CAJ | 학술논문

探索以包涵体形式表达的重组蛋氨酸裂解酶的纯化、复性方法,并对其活性进行检测.将阴道毛滴虫蛋氨酸裂解酶重组表达载体PET-15b-mgl1转化大肠杆菌BL21,经异丙基-β-D-硫代半乳糖苷(IPTG)诱导,并对表达条件进行优化,获得大量表达.通过对包涵体的纯化及复性研究,检测重组蛋氨酸裂解酶的免疫活性及酶活性.Western blotting结果表明,重组蛋氨酸裂解酶免疫小鼠制备的多抗可以和从阴道毛滴虫中提取的天然蛋氨酸裂解酶发生特异性反应.活性检测结果显示复性蛋氨酸裂解酶有活性,复性效率达到25%左右.以包涵体形式表达的蛋氨酸裂解酶经变性、纯化及复性后,获得大量有活性的酶,为深入了解其结构、功能及酶学性质,开展其在临床检测中的应用研究奠定基础.
탐색이포함체형식표체적중조단안산렬해매적순화、복성방법,병대기활성진행검측.장음도모적충단안산렬해매중조표체재체PET-15b-mgl1전화대장간균BL21,경이병기-β-D-류대반유당감(IPTG)유도,병대표체조건진행우화,획득대량표체.통과대포함체적순화급복성연구,검측중조단안산렬해매적면역활성급매활성.Western blotting결과표명,중조단안산렬해매면역소서제비적다항가이화종음도모적충중제취적천연단안산렬해매발생특이성반응.활성검측결과현시복성단안산렬해매유활성,복성효솔체도25%좌우.이포함체형식표체적단안산렬해매경변성、순화급복성후,획득대량유활성적매,위심입료해기결구、공능급매학성질,개전기재림상검측중적응용연구전정기출.
It was to study the purification, renaturation and activity of recombinant Methionine γ-Lyase expressed as inclusion body in E.coli.The E.coli BL21 transformed with PET-15b-mgl1 was cultured and induced by IPTG.The best expression condition of recombinant Methionine γ-Lyase was decided by screening.The purification and renaturation of recombinant Methionine γ-Lyase were explored and the activity was tested. It was proved by Western blotting that the antiserum of Mice immunized with purified Methionine γ-Lyase could specially bind to wild Methionine γ-Lyase purified from Trichomonas vaginalis.Renatured Methionine γ-Lyase with 25% biological activity was prepared after being dissolved, purified and renatured.The results indicate that recombinant MGL expressed as inclusion body retained satisfactory activity after renaturation,suggesting potential efficacy in the research on structure, function, character and clinical trials.