Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin
Defensins are cationic antimicrobial peptides with a broad range of activities against bacteria and fungi. In the present study, the entire coding sequence of codon-optimized Bovine Neutrophil β-Defensin 2 (BNBD2) was designed and placed upstream of Trx coding sequence into the pET-48b (+) vector. F...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2016-09-01
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| Series: | Journal of Advanced Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2090123215001277 |
| _version_ | 1818541076903886848 |
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| author | Shahrzad Aghaei Behnaz Saffar Kamran Ghaedi Mohsen Mobini Dehkordi |
| author_facet | Shahrzad Aghaei Behnaz Saffar Kamran Ghaedi Mohsen Mobini Dehkordi |
| author_sort | Shahrzad Aghaei |
| collection | DOAJ |
| description | Defensins are cationic antimicrobial peptides with a broad range of activities against bacteria and fungi. In the present study, the entire coding sequence of codon-optimized Bovine Neutrophil β-Defensin 2 (BNBD2) was designed and placed upstream of Trx coding sequence into the pET-48b (+) vector. Furthermore, the codon-optimized pelB signal sequences were also added to the upstream of BNBD2 for periplasmic localization. The periplasmic sorting of recombinant β-Defensin 2 was evaluated by osmotic shock and SDS–PAGE on the released proteins. Moreover, the expression of BNBD2-Trx fusion protein was confirmed by the Western blotting technique. Next, the purification of recombinant protein was achieved by Ni++ affinity chromatography. BNBD2 was also separated from Trx by chemical cleavage with formic acid. Finally, both of the antibacterial and antifungal activities of the purified protein were examined. Overall, the results indicated successful periplasmic production of BNBD2 protein, which showed antifungal activity against some of Aspergillus species as well as the antibacterial activity, expressed as successfully suppressed growth of Escherichia coli and Staphylococcus aureus. |
| first_indexed | 2024-12-11T22:03:49Z |
| format | Article |
| id | doaj.art-e12ee31a8b7d4d03b74630ed31711316 |
| institution | Directory Open Access Journal |
| issn | 2090-1232 2090-1224 |
| language | English |
| last_indexed | 2024-12-11T22:03:49Z |
| publishDate | 2016-09-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Advanced Research |
| spelling | doaj.art-e12ee31a8b7d4d03b74630ed317113162022-12-22T00:49:02ZengElsevierJournal of Advanced Research2090-12322090-12242016-09-017581582110.1016/j.jare.2015.12.003Functional analysis of recombinant codon-optimized bovine neutrophil β-defensinShahrzad Aghaei0Behnaz Saffar1Kamran Ghaedi2Mohsen Mobini Dehkordi3Department of Genetics, Faculty of Sciences, Shahrekord University, Shahrekord, IranDepartment of Genetics, Faculty of Sciences, Shahrekord University, Shahrekord, IranDepartment of Biology, Faculty of Sciences, University of Isfahan, Isfahan, IranDepartment of Genetics, Faculty of Sciences, Shahrekord University, Shahrekord, IranDefensins are cationic antimicrobial peptides with a broad range of activities against bacteria and fungi. In the present study, the entire coding sequence of codon-optimized Bovine Neutrophil β-Defensin 2 (BNBD2) was designed and placed upstream of Trx coding sequence into the pET-48b (+) vector. Furthermore, the codon-optimized pelB signal sequences were also added to the upstream of BNBD2 for periplasmic localization. The periplasmic sorting of recombinant β-Defensin 2 was evaluated by osmotic shock and SDS–PAGE on the released proteins. Moreover, the expression of BNBD2-Trx fusion protein was confirmed by the Western blotting technique. Next, the purification of recombinant protein was achieved by Ni++ affinity chromatography. BNBD2 was also separated from Trx by chemical cleavage with formic acid. Finally, both of the antibacterial and antifungal activities of the purified protein were examined. Overall, the results indicated successful periplasmic production of BNBD2 protein, which showed antifungal activity against some of Aspergillus species as well as the antibacterial activity, expressed as successfully suppressed growth of Escherichia coli and Staphylococcus aureus.http://www.sciencedirect.com/science/article/pii/S2090123215001277Antimicrobial peptidesBovine neutrophil beta Defensin 2Codon optimizationPeriplasmic expression |
| spellingShingle | Shahrzad Aghaei Behnaz Saffar Kamran Ghaedi Mohsen Mobini Dehkordi Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin Journal of Advanced Research Antimicrobial peptides Bovine neutrophil beta Defensin 2 Codon optimization Periplasmic expression |
| title | Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin |
| title_full | Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin |
| title_fullStr | Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin |
| title_full_unstemmed | Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin |
| title_short | Functional analysis of recombinant codon-optimized bovine neutrophil β-defensin |
| title_sort | functional analysis of recombinant codon optimized bovine neutrophil β defensin |
| topic | Antimicrobial peptides Bovine neutrophil beta Defensin 2 Codon optimization Periplasmic expression |
| url | http://www.sciencedirect.com/science/article/pii/S2090123215001277 |
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