一株产脂肪酶海洋细菌的筛选、鉴定及其酶学性质

Screening,Identification of A Marine-derived Bacterium Producing Lipase and Its Enzymatic Properties

【目的】研究海洋环境中产脂肪酶菌株的形态、生理生化特性及其所产胞外脂肪酶的酶学性质。【方法】以三丁酸甘油酯作为唯一碳源,利用平板涂布法从黄海胶州湾海泥中筛选到一株产脂肪酶的菌株,对其进行形态、生理生化和分子水平的鉴定,并对其产生的脂肪酶的部分酶学性质进行研究。【结果】产脂肪酶的菌株为薄壁芽孢杆菌属(Gracilibacillus)成员,命名为Gracilibacillus sp.DN-X7。酶学性质研究表明:对该菌所产脂肪酶的最适作用温度、pH和NaCl质量浓度分别为60℃、pH 9.0和30g/L。稳定性试验表明,该酶具有良好的耐热、耐碱和盐稳定性。甘氨酸甜菜碱的存在能够显著提高酶的热稳定性,60℃保温30min酶活力仍保持46%以上。底物特异性试验结果表明DN-X7菌株所产酯裂解酶是脂肪酶,能够分解不同种类的甘油酯和植物油。【结论】分离得到了一株能够产耐热、耐盐的碱性脂肪酶的海洋细菌。

[Objective]A marine-derived bacteria producing extracellular lipase was isolated and i-dentified.The enzymatic properties of the bacteria-produced lipases were characterized.[Method]In this study,tributyrin was used as the sole carbon source.Serial dilution technique was used to isolate microorganisms from marine sediment samples in Jiaozhou Bay of the Yellow Sea on solid agar plates.One of the isolates which produce the highest level of lipase was identified based on morphological,physio-biochemical characteristics and 16S rRNA gene sequence analysis.Moreo-ver,some enzymatic properties of the bacteria-produced lipases were characterized.[Results]The marine-derived bacterium which produce extracellular lipase belonged to Gracilibacillus genus and was designated as Gracilibacillus sp.DN-X7.The result of enzymatic properties showed that the optimal temperature,pH and NaCl concentration of producing lipases were 60 ℃,9.0 and 3%, respectively.The enzyme was adapted to high temperature （〈50 ℃）,alkaline condition （7.0 -12.0）and a wide range of NaCl concentrations.Addition of glycine betaine significantly improved the themostability of the enzyme.The enzyme retained more than 50% of its maximum activity＂nbsp;after 1 h at 60 ℃ in the presence of glycine betaine.Tests of substrate specificity revealed that it was lipase,rather than esterase.The lipase can hydrolyze different kinds of triglycerides and plant oil.[Conclusion]A bacterium that can produce salt-tolerant,thermostable and alkaline li-pases was isolated.