目的:探讨包涵体蛋白的表达条件及纯化方法.方法:将PQE30-Cystatin转化大肠杆菌M15,通过IPTG诱导蛋白表达.为了能够获得可溶的且有活性的Cystatin C(Cys C),本文摸索了Cys C包涵体的复性条件及蛋白表达条件.结果:改变缓冲体系pH值及离子强度等多种方法均不能减少蛋白在复性过程中的沉淀;同时,改变蛋白表达的OD600、培养基pH值、表达时间等均未得到可溶性的表达蛋白;而改变诱导蛋白表达所用的IPTG浓度后发现在低浓度(0.005mM)的IPTG诱导条件下,该蛋白有少量的表达,但是表达量明显低于1mM IPTG的诱导条件.结论:对于包涵体蛋白进行方法摸索的过程中,既要对包涵体蛋白本身进行研究,更要对蛋白表达条件进行进一步的优化,以表达可溶的蛋白.%Objective To study the optimal conditions for expression of inclusion body protein and method of purification of the protein. Methods PQE30-Cystatin C was transformed into E-Coli M15 and protein expression was induced with IPTG. Options for better renaturation of the obtained inclusion body protein were studied. Results We tried various ways including changes of buffer system pH and ionic strength, as well as OD 600 for protein expression, culture media pH, expression time ……but failed to obtain soluble expression protein. However, we lowered the concentration of IPTG solution from 1mmol/L to 0.005mmol/L and finally obtained a slight amount of expressed soluble fusion Cystatin C protein. Conclusion Presently, the amount of soluble protein obtained with low concentration of IPTG is too little for practical use and further study is needed.
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