Research on Lipolytic Enzymes from Bacillus licheniformis CP-16

LI Shuang, ZHANG Tieying, LIAO Chaoyong, LIANG Zhengwen, MA Xiafei, LIU Junli

doi:10.3969/j.issn.1006-267x.2017.11.026

Chinese Journal of Animal Nutrition >

2017 , Vol. 29 >Issue 11: 4048 - 4057

DOI: https://doi.org/10.3969/j.issn.1006-267x.2017.11.026

Molecular Nutrition

Research on Lipolytic Enzymes from Bacillus licheniformis CP-16

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1. State Key Laboratory of Animal Nutrition, Institute of Animal Science, Chinese Academy of Agricultural Science, Beijing 100193, China;
2. College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China

Received date: 2017-05-02

Online published: 2017-10-31

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Abstract

In order to explore the influence of lipolytic enzyme on feather degradation, the lipolytic enzyme gene of Bacillus licheniformis CP-16 was cloned and heterologous expressed in this research. The target gene of recombinant enzyme L-4, which was used Bacillus licheniformis CP-16 genome DNA as a template, amplified lipid hydrolase gene, and then transferred into Escherichia coli for expression of the targeted gene. The optimum pH, pH stability, temperature, temperature stability and effects of organic solvents and metal ions on relative activity of the recombinant enzyme L-4 were determined, and its application on keratin K hydrolyzed natural feather keratin was also investigated. The results showed that the length of lipid hydrolase gene was 747 bp, encoded 248 amino acids, the recombinant enzyme L-4 was successfully expressed in Escherichia coli, and the molecular weight was about 28.3 ku, the esterase activity was 0.41 U/mL, the optimum pH was 6.5, the optimum temperature was 50℃. The relative activity of recombinant enzyme L-4 kept above 80% treated with 30 min under pH 6.5 to 9.5 condition, and the relative activity kept above 70% treated with 30 min under below 50℃ condition. The ferrous iron (Fe²⁺), sodion (Na⁺),manganese ion (Mn²⁺) and calcium ion (Ca²⁺) had an stimulative effect on the relative activity of recombinant enzyme L-4, while the barium ion (Ba²⁺), zinc ion (Zn²⁺), copper ion (Cu²⁺) and nickel ion (Ni²⁺) had an disincentive effect on the relative activity of recombinant enzyme L-4. When the concentration of organic solvent was 30%, the relative activity of recombinant enzyme L-4 kept 85% and 97% in methanol and dimethyl sulfoxide solutions, the relative activity kept above 45% in acetone and ethanol solutions, the relative activity kept less than 20% in isopropanol solution, while the relative activity was complete loss in acetonitrile solution. The pretreatment of natural feather substrates with recombinant enzyme L-4 promoted the hydrolysis of keratinase K to the substrate, and the promotion rate is 4.32%. In conclusion, lipolytic enzymes can degrade feather surface lipids and it may play a role in promoting keratin hydrolysis of feather keratin.

Key words： Bacillus licheniformis; lipolytic enzymes; keratin; feather lipid; cloning and expression

Cite this article

LI Shuang, ZHANG Tieying, LIAO Chaoyong, LIANG Zhengwen, MA Xiafei, LIU Junli . Research on Lipolytic Enzymes from Bacillus licheniformis CP-16[J]. Chinese Journal of Animal Nutrition, 2017 , 29(11) : 4048 -4057 . DOI: 10.3969/j.issn.1006-267x.2017.11.026

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