以产腈水解酶重组大肠杆菌E.coli BL21(DE3)/pET-28b-NIT为出发菌株,在摇瓶培养的基础上,建立5 L发酵罐上重组大肠杆菌产腈水解酶的高密度发酵工艺。通过分批补料培养工艺优化,表明碳、氮源的浓度以及诱导剂的浓度和诱导时机是影响菌体生长和酶活的重要因素。在优化条件下,对E.coli BL21(DE3)/pET-28b-NIT进行14 h的分批补料培养,菌体细胞浓度为13.7 gDCW/L,较优化前提高了4.3倍;腈水解酶体积酶活为36 990 U/L,较优化前提高了5.1倍。

Methionine is not only an essential amino acid needed for human health,also an important feed additive.The recombinant E.coli BL21(DE3) /pET-28b-NIT producing nitrilase was investigated.Based on the data obtained from the cultivation using shake flask,the fed-batch fermentation strategy for producing nitrilase by high cell density cultivation of recombinant E.coli in the 5-Liter fermentor was optimized and constructed.It indicated that the concentration of carbon source,nitrogen source and inducer were the important factors.Under the optimized conditions of 14 h fed-batch fermentation,the cell mass concentration and the specific activity reached 13.7 gDCW/L and 36 990 U/L,which was 4.3 and 5.1 times higher than previous optimization,respectively.