大肠杆菌不耐热肠毒素(LT)无毒突变体mLT63在大肠杆菌中的融合表达及纯化

目的 在大肠杆菌中表达并纯化大肠杆菌不耐热肠毒素无毒突变体mLT63融合蛋白，并探索其免疫反应性。方法采用不同的诱导温度，IPTG浓度，葡萄糖含量，pH值以及不同的培养基优化重组菌TB1（pMAL-c2X-mlt）的表达条件，表达产物经超声粉碎,超声上清经初步盐析、应用直链淀粉亲和层析柱纯化，Western blot鉴定纯化产物的免疫学反应性。结果 融合蛋白在诱导表达产物中的含量达到16．11％,纯化产物的纯度达到72．58％。纯化产物能够被抗CT血清所识别。结论 成功表达并纯化出了大肠杆菌不耐热肠毒素无毒突变体mLT63，获得了纯度较高并具有良好免疫反应性的融合蛋白。

Expression and purification of the fusion protein of the avirulent mutant mLT63 from heat-labile enterotoxin of E.coli

To express and purify the fusion protein of the avirulent mutant mLT63 from heat-labile enterotoxin of E.coli and to determine its immunoreactivity with specific antibodies,different conditions for expression,such as induction temperature,concentration of IPTG,content of glucose,pH value and different culture media were investigated in order to optimize the expression of the recombinant strain TB1(pMAL-c2X-mlt).The expressed product was purified through amylose pre-packed column after treatment with ultrasonic disintegrator and salting out of the supernatant after ultrasonication.SDS-PAGE was used for the detection of the expressed product and Western blot was employed to determine the immunoreactivity of mLT63 with rabbit anti-cholera toxin(CT) serum.It was demonstrated that the content of the fusion protein from recombinant mLT63 was found to be 16.11% with a purity of 72.58% after purification.In addition,this fusion protein could be recognized by rabbit anti-CT serum.with high immunoreactivity.In the present investigation,the fusion protein of the avirulent mutant mLT63 from heat-labile enterotoxin of E.coli was successfully obtained.