药物生物技术
藥物生物技術
약물생물기술
PHARMACEUTICAL BIOTECHNOLOGY
2001年
1期
22-25
,共4页
韩照中%王恒樑%冯尔玲%苏国富%黄翠芬
韓照中%王恆樑%馮爾玲%囌國富%黃翠芬
한조중%왕항량%풍이령%소국부%황취분
L-天门冬酰胺酶II%表达%纯化
L-天門鼕酰胺酶II%錶達%純化
L-천문동선알매II%표체%순화
L-asparaginase II%Expression%Purification
依据发表的大肠杆菌天门冬酰胺酶II基因序列，合成引物，利用PCR技术获得天门冬酰胺酶II的结构基因，插入高效表达载体pBV220，并转化大肠杆菌菌株DH5α，获得高效表达天门冬酰胺酶II的基因工程菌株。通过对包涵体的提取与纯化，结合离子交换层析获得天门冬酰胺酶II的蛋白质纯品。检测结果表明得到的天门冬酰胺酶II具有较高的比活性，与商品化的同类产品比活相近。
依據髮錶的大腸桿菌天門鼕酰胺酶II基因序列，閤成引物，利用PCR技術穫得天門鼕酰胺酶II的結構基因，插入高效錶達載體pBV220，併轉化大腸桿菌菌株DH5α，穫得高效錶達天門鼕酰胺酶II的基因工程菌株。通過對包涵體的提取與純化，結閤離子交換層析穫得天門鼕酰胺酶II的蛋白質純品。檢測結果錶明得到的天門鼕酰胺酶II具有較高的比活性，與商品化的同類產品比活相近。
의거발표적대장간균천문동선알매II기인서렬，합성인물，이용PCR기술획득천문동선알매II적결구기인，삽입고효표체재체pBV220，병전화대장간균균주DH5α，획득고효표체천문동선알매II적기인공정균주。통과대포함체적제취여순화，결합리자교환층석획득천문동선알매II적단백질순품。검측결과표명득도적천문동선알매II구유교고적비활성，여상품화적동류산품비활상근。
A pair of primers was designed and synthesized based on publishedsequence, and used to amplify the structural gene fragment of L-asparaginase II of E.coli by polymerase chain reactions(PCR) . A recombinant plasmid was const ructed by placing L-asparaginase II gene under lamda PLPR promotor. The recombinant bacterium harboring the recombinant plasmid expressed L-asparaginase II in high level as inclusion bodies. After isolation ,washing of inclusion bodies, denatur ization-renaturization and one-stepion-exchanging chromatography ,active L -asp araginase II was obtained ,which has activity in vitro similar to the commer cialized products.
