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| A novel lipase gene of Aspergillus niger C112 cloning, and heterologous expression optimization |
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| DOI：10.19902/j.cnki.zgyz.1003-7969.240617 |
| KeyWord:Aspergillus niger C112 lipase Pichia pastoris bioinformatics cloning |
| FundProject:木本油料资源利用全国重点实验室基金项目（GZKF202107）；湖南省研究生科研创新项目（CX20220722） |
| Author Name | Affiliation | | XU Pengjun1,2, ZENG Baiquan1,2, XIAO Zhihong3, LI Meiqun1,3, ZHANG Xuan1,3,
LI Peiwang3, ZHAO Xitong1, YAN Li′an1 | 1.College of Life Science and Technology, Central South University of Forestry and Technology,
Changsha 410004, China
2.State Key Laboratory of Utilization of Woody Oil Resource, Central
South University of Forestry and Technology, Changsha 410004, China
3.State Key Laboratory of
Utilization of Woody Oil Resource, Hunan Academy of Forestry, Changsha 410004, China |
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| Abstract: |
| To provide a reference for the modification of lipase and its industrial application, the lipase gene was amplified by PCR based on the Aspergillus niger C112 strain, and the gene was analyzed by bioinformatics. The recombinant Pichia pastoris GS115 strain was constructed by genetic engineering techniques,and the lipase induced by the recombinant strain was purified and its enzymatic properties were analyzed. In addition, the lipase production conditions with the recombinant strain were optimized through single factor experiment and response surface methodology. The results indicated that the length of the lipase gene fragment was 894 bp, and its theoretical molecular weight was 31.79 kDa. A new pentapeptide RHSLG was found in its amino acid sequence. The optimal reaction temperature and pH of the recombinant lipase were 40 ℃ and 4, respectively, which showed good thermal stability and pH stability. The optimal production conditions of the recombinant strain were 10.50 mL of YNB added in 100 mL BMMY medium, inoculation amount 4.3%, adding 1% methanol every 24 h, and a culture time of 2.8 d. Under the optimal conditions, the enzyme activity of the recombinant lipase reached 78.92 U/mL, which was 1.79 times that before optimization. In summary, the recombinant lipase after cloning and heterologous expression has strong stability and high enzyme activity. |
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