Volume 10, Issue 39 (6-2020)                   NCMBJ 2020, 10(39): 91-102 | Back to browse issues page

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Mirzaei M, Rastegar Shariat Panahi M, Ghahremani E, Rezae E, Seid Moradi R, Farnoosh G et al . Improving the activity and stability of OPH enzyme using DNA Shuffling method. NCMBJ 2020; 10 (39) :91-102
URL: http://ncmbjpiau.ir/article-1-1294-en.html
Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
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Improving the activity and stability of OPH enzyme using DNA Shuffling method
Morteza Mirzaei 1, Mona Rastegar Shariat Panahi 2, Enayat Ghahremani 1, Ehsan Rezae 3, Rezvan Seid Moradi 1, Gholamreza Farnoosh1, Ali Mohammad Latifi 1,*
1- Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
2- Department of Biochemistry, Payame Noor University, Tehran, Iran
3- Molecular Biology Research Center, Systems biology and poisonings Institute, Baqiyatallah University of Medical   Sciences, Tehran, Iran
Abstract
Aim and background: One of the effective methods for protein engineering, especially for improving the capabilities of industrial strains, is the genome shuffling method. This study is based on biocatalytic engineering techniques and DNA shuffling methods.
Materials and Methods: Pseudomonas aeruginosa wild strain was prepared under the access number JQ917006.1. Mutant strains were evaluated by DES method and compatibility with diazinon concentration. After the protoplast was prepared, the genome was mutated from the mutant library. Fused protoplasts were evaluated for activity.
Results: The activities related to IR1.G1, IR1.D8, IR1.D4 and IR1.D5 strains are 0.234 U/ml, 0.1 U/ml, 0.098 U/ml and 0.066 U/ml, respectively, and IRL1.F2, IRL1.F3 and IRL1.F1 strains have activities of 0.541 mg/L, 0.523 mg/L and 0.509 mg/L, respectively.
 
Conclusion: The results of evaluation of the first generation of genome shuffling (first round of protoplast fusion) showed that the shuffled strains that were able to grow in the presence of toxin (3000 ml/L diazinon concentration) had better activity than the obtained strain through both methods (toxin concentration gradient and DES method).
Keywords: Genome shuffling, improved strains, protein engineering, diazinon

 
 
 
 
 
 
 
Type of Study: Research Article | Subject: Cellular and molecular
Received: 2020/06/27 | Accepted: 2020/06/27 | Published: 2020/06/27

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