一株产耐有机溶剂脂肪酶伯克霍尔德氏菌的分子鉴定及酶学性质分析Molecular identification and enzymatic properties of an organic solvent tolerant lipase-producing Burkholderia sp. strain

Abstract

为筛选具有潜在应用价值的产脂肪酶菌株，采用三丁酸甘油酯固体培养基从黄山松树林土壤中筛选产脂肪酶菌株，利用16S rDNA测序对筛选的菌株进行鉴定，并对其产脂肪酶酶活力参数以及酶学性质进行分析。结果表明：筛选的产脂肪酶菌株HSU-7为伯克霍尔德氏菌(Burkholderia sp. HSU-7)；菌株HSU-7所产脂肪酶的最适反应温度为45 ℃，最适pH为5，为一种耐酸性的中温脂肪酶；菌株HSU-7发酵5 d，所产脂肪酶酶活力最大，为72.22 U/mL。同时，K+、Ca2+和Mg2+对菌株HSU-7所产脂肪酶酶活力具有显著的促进作用，而Ni2+、Cu2+、Fe3+和Zn2+具有一定的抑制作用，该脂肪酶可耐受有机溶剂甲醇、乙醇、异丙醇和二甲基亚砜，但Triton X-100可显著抑制其酶活力；此外，该脂肪酶能耐受65 ℃的高温。综上，菌株HSU-7可高产脂肪酶，且该脂肪酶有较强的热稳定性，可耐受多种有机溶剂，具有很好的工业应用价值。In order to screen the lipase-producing strain with potential applications, the solid medium containing tributyrin was used to screen the lipase-producing strain from the soil of pine forest in Huangshan. Then the lipase-producing strain were identified by 16S rDNA sequencing and the enzymatic activity parameters and properties of the lipase produced were determined. The results showed that the lipase-producing strain HSU-7 selected was identified as Burkholderia sp.. The optimum reaction temperature and pH for the lipase produced by the strain HSU-7 were 45 ℃ and 5, respectively, revealing that it was an acid-resistant and medium-temperature lipase. The enzymatic activity of lipase produced by strain HSU-7 was the highest at 72.22 U/mL after 5 d of fermentation. Meanwhile, K+, Ca2+, and Mg2+ significantly promoted the enzymatic activity of the lipase produced by strain HSU-7, while Ni2+, Cu2+, Fe3+, and Zn2+ had some inhibition effects on the enzymatic activity. The lipase could tolerate the organic solvents methanol, ethanol, isopropanol and dimethyl sulfoxide, but Triton X-100 significantly inhibited the activity of the lipase. In addition, the lipase could tolerate 65 ℃. In summary, the strain HSU-7 can produce lipase with high yield, and the lipase has strong thermal stability and can tolerate a variety of organic solvents, so the strain has good industrial application value