高产磷脂酶D的蜡样芽孢杆菌诱变筛选及发酵条件优化  

Mutation screening and fermentation condition optimization of Bacillus cereus for high production of phospholipase D

作　　者：王天一 刘欢 王芳[1] 邓利[1] WANG TianYi;LIU Huan;WANG Fang;DENG Li(College of Life Science and Technology,Beijing University of Chemical Technology,Beijing 100029,China)

机构地区：[1]北京化工大学生命科学与技术学院,北京100029

出　　处：《北京化工大学学报（自然科学版）》2023年第5期71-79,共9页Journal of Beijing University of Chemical Technology(Natural Science Edition)

基　　金：国家自然科学基金(21978019/21978020)。

摘　　要：蜡样芽孢杆菌(Bacillus cereus)是一种天然的磷脂酶D(PLD)生产菌株,为了提高PLD产量,以野生型蜡样芽孢杆菌(B.cereus CICC 20551)为出发菌,使用紫外线和硫酸二乙酯对其进行复合诱变,通过建立的PLD活性高效筛选方法,从大量突变样本中筛选得到一株高产PLD的蜡样芽孢杆菌S6-UD46。将该菌株摇瓶发酵48 h,产酶量达到0.52 U/mL,相比原始菌株(0.32 U/mL)提高了62.5%。对诱导产酶条件与培养基成分进行了优化,确定最佳发酵条件为:以20 g/L牛肉膏为氮源,10 g/L蔗糖为碳源,在600 nm波长下的光密度(OD600)达到4左右时加入40 g/L卵磷脂(磷脂酰胆碱含量>90%)对突变菌株S6-UD46进行诱导产酶,在37℃、200 r/min下培养48 h,PLD酶活性可达3.22 U/mL,是优化前的6.19倍。Bacillus cereus is a natural phospholipase D(PLD)producing strain.In order to improve the yield of PLD,the wild-type B.cereus CICC 20551 was taken as the starting strain and subjected to complex mutagenesis with ultraviolet light and diethyl sulfate.A strain of B.cereus S6-UD46 with high PLD activity was screened from a large number of mutant samples by the established PLD activity efficient screening method.The enzyme production of the strain reached 0.52 U/mL after shaking flask fermentation for 48 h,which was 62.5%higher than that of the original strain(0.32 U/mL).The optimal fermentation conditions were determined as follows:using 20 g/L beef extract as nitrogen source,10 g/L sucrose as carbon source,40 g/L lecithin(phosphatidylcholine content>90%)was added when the optical density at 600 nm wavelength reached about 4 to induce the enzyme production by mutant strain S6-UD46.After incubation at 37℃and 200 r/min for 48 h,the PLD activity was 3.22 U/mL,which is 6.19 times the value before optimization.

关 键 词：磷脂酶D 蜡样芽孢杆菌 诱变筛选 发酵条件优化 

分 类 号：Q815[生物学—生物工程]