Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot
Background: Pseudomonas aeruginosa is a common Gram-negative, rod-shaped bacterium that has a unique genome that allowed the bacteria to produce various enzymes and proteins. Azurin and arginine deiminase are low molecular weight proteins that produced by P. aeruginosa. These proteins can be used al...
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doaj-945fcdc602c845a1bf0bc6558b2864462020-11-25T03:44:08ZengShahid Beheshti University of Medical SciencesNovelty in Biomedicine2345-39072020-06-0182637010.22037/nbm.v1i1.2646113036Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western BlotMina Hadi0Mona Ghazi1Parvaneh Saffarian2Bahareh Hajikhani3Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.Department of Microbiology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.Department of biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.Department of Microbiology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.Background: Pseudomonas aeruginosa is a common Gram-negative, rod-shaped bacterium that has a unique genome that allowed the bacteria to produce various enzymes and proteins. Azurin and arginine deiminase are low molecular weight proteins that produced by P. aeruginosa. These proteins can be used alone or in combination together in order to become effective in cancer therapy or inhibiting of metastasis. This study aimed to design, express and purify the Azurin and Arginine Deiminease recombinant fusion protein. Materials and Methods: The sequences of Azurin and arginine deiminase from P. aeruginosa were studied for synthesis in a pET28a vector. The recombinant plasmid was transfected into the E.coli BL-21 strain and expression was induced by isopropyl-β-D-thio galactopyranoside (IPTG). The fusion protein expression was evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Purification of the recombinant product was performed by the Ni-NTA chromatography column, obtained product analysis with SDS-PAGE and Western blot technique. Results: Cloning was confirmed by observing bands from the enzyme digestion. The protein band with a molecular weight of 65 kDa on the SDS-PAGE gel was an indication of the correct expression of the protein. The single-band of this recombinant protein was confirmed by the western blot technique. Conclusion: In this study, due to the successful production of arginine deiminase-azurin fusion protein, and considering the separate anti-cancer properties of these compounds, which have been reported in previous studies, it is suggested that immunological assessments and effects of this fusion protein in different cancerous cell line be investigated.http://journals.sbmu.ac.ir/nbm/article/view/26461pseudomonas aeruginosaazurinarginine deiminasefusion protein. |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Mina Hadi Mona Ghazi Parvaneh Saffarian Bahareh Hajikhani |
spellingShingle |
Mina Hadi Mona Ghazi Parvaneh Saffarian Bahareh Hajikhani Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot Novelty in Biomedicine pseudomonas aeruginosa azurin arginine deiminase fusion protein. |
author_facet |
Mina Hadi Mona Ghazi Parvaneh Saffarian Bahareh Hajikhani |
author_sort |
Mina Hadi |
title |
Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot |
title_short |
Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot |
title_full |
Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot |
title_fullStr |
Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot |
title_full_unstemmed |
Design and Production of Arginine Deiminase-Azurin Recombinant Fusion Protein from Pseudomonas aeruginosa and its Confirmation by Western Blot |
title_sort |
design and production of arginine deiminase-azurin recombinant fusion protein from pseudomonas aeruginosa and its confirmation by western blot |
publisher |
Shahid Beheshti University of Medical Sciences |
series |
Novelty in Biomedicine |
issn |
2345-3907 |
publishDate |
2020-06-01 |
description |
Background: Pseudomonas aeruginosa is a common Gram-negative, rod-shaped bacterium that has a unique genome that allowed the bacteria to produce various enzymes and proteins. Azurin and arginine deiminase are low molecular weight proteins that produced by P. aeruginosa. These proteins can be used alone or in combination together in order to become effective in cancer therapy or inhibiting of metastasis. This study aimed to design, express and purify the Azurin and Arginine Deiminease recombinant fusion protein.
Materials and Methods: The sequences of Azurin and arginine deiminase from P. aeruginosa were studied for synthesis in a pET28a vector. The recombinant plasmid was transfected into the E.coli BL-21 strain and expression was induced by isopropyl-β-D-thio galactopyranoside (IPTG). The fusion protein expression was evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Purification of the recombinant product was performed by the Ni-NTA chromatography column, obtained product analysis with SDS-PAGE and Western blot technique.
Results: Cloning was confirmed by observing bands from the enzyme digestion. The protein band with a molecular weight of 65 kDa on the SDS-PAGE gel was an indication of the correct expression of the protein. The single-band of this recombinant protein was confirmed by the western blot technique.
Conclusion: In this study, due to the successful production of arginine deiminase-azurin fusion protein, and considering the separate anti-cancer properties of these compounds, which have been reported in previous studies, it is suggested that immunological assessments and effects of this fusion protein in different cancerous cell line be investigated. |
topic |
pseudomonas aeruginosa azurin arginine deiminase fusion protein. |
url |
http://journals.sbmu.ac.ir/nbm/article/view/26461 |
work_keys_str_mv |
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