We found a match
Your institution may have access to this item. Find your institution then sign in to continue.
- Title
阴沟肠杆菌WJ-1中甲氰菊酯降解酶基因pytZ的克隆、表达和重组酶特性研究.
- Authors
阎 华; 雷 婷; 汤尚文; 于 博; 黄升谋; 李云捷; 吴进菊; 李玉奇
- Abstract
[Objective] The pesticide degrading enzymes were generally more tolerant to abnormal environment and had wider degradation spectrum than pesticide degrading bacteria. In order to obtain high yield and high quality pesticide degrading enzyme, based on the previous acquisition of Enterobacter cloacae WJ-1 and the confirmation of its pesticide degrading enzyme gene pytZ in the laboratory, the pesticide degrading enzyme gene pytZ of E. cloacae WJ-1 was recombined and expressed in the recipient strain Escherichia coli BUl. And the physio- logical and biochemical characteristics of the recombinant enzyme were studied. [Method] The pesticide degrading enzyme gene pytZ of E. cloacae WJ-1 was amplified by PCR, and the recombinant plasmid pMD19 was constructed. Then the recombinant plasmid pMD19 was ligated with pET-24a(K +) expression vector to construct the genetically engineered strain Escherichia coli BL21/pET-24a (K +)-pytZ. The re-combinant bacteria were cultured in shake flask. The kinetic parameters of recombinant enzyme were studied by using different temperature, pH value and IPTG concentration. [ Result]The E. coli BL21/pET-24a(K +)-pytZ was obtained. At 30 "C and IPTG induction concentration of 0. 8 mmol/L, the highest yield of pesticide degrading enzyme was obtained after 48 hours in shake flask culture, and the enzyme activity reached 15. 3 U/ mL The optimal reaction temperature of the recombinant enzyme was 35 "C, 25 - 45 "C with good thermal stability; The results showed that the optimal pH value of the enzyme was 8. 0, pH 7. 0 - 9. 0, and the acid-base stability was good; The Michaelis constant Km was 0. 076 mmol/L, and the maximum reaction rate Vmax was 3. 023 mmol/( L · min). [Conclusion] It was found that more fenpropathrin degrading enzymes can be produced by Escherichia coli BL21/pET-24a (K +) -pytZ, with higher purity, wider adaptability to temperature and pH, and higher degradation efficiency.
- Subjects
ENTEROBACTER cloacae; ESCHERICHIA coli; THERMAL stability; PESTICIDES; ENZYMES; BOTTLES
- Publication
Southwest China Journal of Agricultural Sciences, 2021, Vol 34, Issue 8, p1762
- ISSN
1001-4829
- Publication type
Article
- DOI
10.16213/j.cnki.scjas.2021.8.025