重组尿酸氧化酶发酵工艺优化

目的:优化并获得重组尿酸氧化酶(rUOX)基因工程大肠杆菌BL21(DE3)/pET-32a-uox高密度发酵的工艺参数。方法:在三角摇瓶中进行培养条件的优化实验,分别考察了pH值、接种量、无机盐、碳源、诱导强度等对工程菌生长和重组蛋白表达的影响,得到了优化的发酵条件;在此基础上放大至NBS BIOFLO 110 14 L发酵罐,通过对诱导时机的优化,利用分批补料发酵的方式,使rUOX在高密度培养的条件下得到高表达。结果:在优化的发酵条件下,菌体密度(D600nm)最终达到50以上,相当于20 g/L干重;可溶性rUOX占菌体总蛋白量的45%,其含量达到3.45 g/L。结论:为规模化制备重组黄曲霉尿酸氧化酶奠定了基础。

Fermentation Process Optimization of Recombinant Urate Oxidase

Objective： To optimize the high density fermentation process of recombinant urate oxidase（rUOX）. Methods： In order to optimize the fermentation conditions of E.coli BL21 （DE3）/pET32auox, flask shaking cul ture was done to get the optimized culture conditions. Firstly, effects of carbon, phosphate, inoculation and pH were investigated; then, the induction conditions were optimized, including induction time, induction duration and inducer concentration of IPTC. Based on these data, fedbatch fermentation was carried out on NBS BIOFLOll0 14 L fermentor to high celldensity culture recombinant E.coli BL21（DE3）/pET32auox. Bacterial were harvest af ter 5 h induction of 0.5 mmol/L IPTG. Results： Under the optimized conditions, the final cell density was more than 50（D~0.m）. The amount of synthesized rUOX was up to 45% of total bacterial proteins, the final yield of puri fied rUOX from each liter of high celldensity culture was about 3.45 g. Conclusion： This study lays a founda tion for production of recombinant rUOX in scale.