耐有机溶剂脂肪酶产生菌的筛选及其粗酶酶学性质

以体积分数2%苯为筛选压力,利用罗丹明B平板显色法和摇瓶发酵法,从采集的花生地土壤样品中分离筛选得到1株中度耐热、耐碱脂肪酶产生菌,编号为H2。通过形态观察、生理生化特性实验及其16S rDNA基因序列对菌种进行鉴定。结果表明,H2菌株与短小芽孢杆菌(Bacillus pumilus)的亲缘关系最紧密。通过研究得到该菌株的摇瓶发酵条件:产酶培养基为:蛋白胨3%、酵母膏1%、NaCl 0.5%、橄榄油1%,pH7.0,摇瓶发酵温度为28℃,摇床转速为180r/min,发酵周期为48～60h。所产脂肪酶在40℃、pH9.0时酶活性最高,对pH值和温度的适应范围较宽,pH6.0～10.0比较稳定,35～50℃具有较高酶活性。

Screening of Organic Solvent Tolerant Lipase-Producing Bacteria and Enzymatic Properties of Crude Lipase

A moderately thermostable and alkali-resistant lipase-producing strain named as H2 was isolated from peanut plantation soil using 2% benzene by Rhodamine B plate assay and shake flask fermentation.It was preliminarily identified based on morphological and physiological characteristics.Meanwhile,the 16S rDNA gene of the strain was cloned and sequenced.The physiological and biochemical characterization and phylogenetic tree based on the 16S rDNA sequence showed that the strain was highly homologous to Bacillus pumilus.The optimal culture medium for lipase production by the strain was composed of 3% peptone,1% yeast extract,0.5% NaCl,1% olive oil,pH 7.0,and the optimal fermentation conditions of temperature,rotational speed and time were 28 ℃,180 r/min and 48-60 h,respectively.The lipase produced under these conditions revealed the highest activity at 40 ℃ and pH 9.0.The lipase was stable over a broad range of pH(6.0-10.0) and temperature(35-50 ℃).