Cloning, Expression and Characterization of the Anticancer Protein Azurin from an Indigenous Strain Pseudomonas aeruginosa SSj
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Azurin is a novel anticancer protein, mainly produced from Pseudomonas aeruginosa and few other Gram-negative bacteria. In this study, azurin gene (azu) from a native Pseudomonas aeruginosa strain SSj was amplified using PCR with specific primers. The azurin gene (418 bp) was sequenced and submitted in Genbank. The PCR amplicon was digested with BamHI and HindIII and inserted into expression vector pET-22b(+). The recombinant protein was expressed in Escherichia coli BL21 (DE3) after induction with IPTG. Protein purification was done using Ni NTA agarose. The SDS-PAGE analysis of purified samples with Coomassie brilliant blue showed a band with an apparent molecular weight of about 14 kDa. Structural characterization of purified protein was done by FTIR, MALDI-TOF and LC–MS/MS analysis.
KeywordsAzurin BL21 (DE3) pET-22b(+) MALDI-TOF LC–MS/MS Anticancer protein
Polymerase chain reaction
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- Ni NTA
Fourier-transform infrared spectroscopy
Matrix assisted laser desorption/ionization time of flight
Liquid chromatography with tandem mass spectrometry
The authors would like to extend their gratitude to Department of Life Sciences, University of Calicut, India, for providing necessary facilities.
This work was carried out in collaboration between both authors. Author DS designed the study and done supervision and manuscript correction. Author MCS managed the literature searches, manuscript work and the experimental process. Both authors read and approved the final manuscript.
This work was supported by Innovative Research Project, University of Calicut.
Compliance with Ethical Standards
Conflict of interest
Authors have no conflict of interest regarding the publication of paper.
- Mohamed MS, Fattah SA, Mostafa HM (2010) Azurin as antitumor protein and its effect on the cancer cell lines. Curr Res J Biol Sci 2(6):396–401Google Scholar
- Sankar R, Siddik S, Abhijit M, Mahitosh M (2011) Azurin synthesis from Pseudomonas Aeruginosa MTCC 2453, properties, induction of reactive oxygen species, and p53 stimulated apoptosis in breast carcinoma cells. J Cancer Sci Ther 03:104–111. https://doi.org/10.4172/1948-5956.1000069 CrossRefGoogle Scholar