骆驼凝乳酶工程菌发酵培养基的优化

Optimization of Fermentation Medium for Genetically Engineered Strain with Camel Chymosin-producing capability

摘要:

凝乳酶可以专一地切割乳中κ-酪蛋白的Phe 105-Met 106之间的肽键，引起蛋白质凝聚，在乳制品加工中应用广泛。目前对牛凝乳酶的研究比较成熟，而对骆驼凝乳酶的研究报道较少。本研究为获得高效骆驼凝乳酶发酵培养基，在单因素实验的基础上，采用Plackett-Burman试验考察培养基中的8个成分对骆驼凝乳酶产量的影响。研究表明葡萄糖、玉米浆、尿素的对产酶影响显著。采用最陡爬坡试验逼近最大响应区间，利用Box-Behnken响应面法对3个主要影响因素进行分析，得到高效发酵培养基配方：葡萄糖86.6 g/L、玉米浆56.9 g/L、酵母粉4 g/L、磷酸二氢钾3 g/L、尿素18 g/L、硫酸锌3 g/L、硫酸氨1 g/L和氯化镁0.3 g/L。在该条件下凝乳酶活达到431.25 U/mL，较优化前的基础培养基（207.03 SU/mL）提高了2.08倍。再利用1L发酵罐进行发酵培养，凝乳酶活性达到530 SU/mL，约为摇瓶培养最高酶活性的1.22倍。

Abstract:

Chymosin can specifically cleave the peptide bond between κ-casein Phe 105 and Met 106 in milk, cause milk-clotting and widely used in dairy products. Bovine chymosi has been deeply studied, but few research on camel chymosin were reported. According to the single-factor experiments design, Plackett-Burman design was used to evaluate the effects of 8 medium components on the production of chymosin to obtain a high-performance fermentation medium. The results showed that glucose, corn syrup, and urea concentrations had significant effects on the chymosin production. Box-Behnken experimental design was employed to optimize the three medium components furtherly. The optimal fermentation medium for antifungal substance production was composed of glucose 86.6 g/L, corn syrup 56.9 g/L, yeast extract 4g/L , KH2PO4 3 g/L, urea 18 g/L, ZnSO4 3 g/L, (NH4)2SO4 1 g/L and MgCl2 0.3 g/L. Under the optimal conditions, the activity of chymosin could reach 431.25 SU/mL, which were 2.08 folds higher than the original medium (207.03 SU/mL). The chymosin activity reached 530 SU/mL after fermentation in an 1Lfermentation tank with exact medium components,being 1.22 folds higher than the chymosin activity obtained by shake flask fermentation.