Abstract
Brucellosis is a disease caused by bacteria belonging to the genus Brucella. It affects cattle, goat, sheep, dog and humans. The serodiagnosis of brucellosis involves detection of antibodies generated against the LPS or whole cell bacterial extracts, however these tests lack sensitivity and specificity. The present study was performed to optimize the culture condition for the production of recombinant Brucella melitensis outer membrane protein 28 kDa protein in E.coli via fed batch fermentation. Expression was induced with 1.5mM isopropyl β thiogalactoside and the expressed recombinant protein was purified using Ni-NTA affinity chromatography. After fed-batch fermentation the dry cell weight of 17.81 g/L and a purified protein yield of 210.10 mg/L was obtained. The purified Brucella melitensis recombinant Omp 28 kDa protein was analyzed through SDS- poly acrylamide gel electrophoresis and western blotting. The obtained recombinant protein was evaluated for its diagnostic application through Indirect ELISA using brucellosis suspected human sera samples. Our results clearly indicate that recombinant Omp28 produced via fed batch fermentation has immense potential as a diagnostic reagent that could be employed in sero monitoring of brucellosis.
Keywords: Fed batch fermentation, brucellosis, diagnosis, outer membrane protein, ELISA
Protein & Peptide Letters
Title:Fed Batch Fermentation and Purification Strategy for High Yield Production of Brucella melitensis Recombinant Omp 28 kDa Protein and its Application in Disease Diagnosis
Volume: 20 Issue: 7
Author(s): Karothia B.S., Athmaram T.N., Thavaselvam D., Kumar Ashu, Sapna Tiwari, Anil K. Singh, Sathyaseelan K. and Gopalan N.
Affiliation:
Keywords: Fed batch fermentation, brucellosis, diagnosis, outer membrane protein, ELISA
Abstract: Brucellosis is a disease caused by bacteria belonging to the genus Brucella. It affects cattle, goat, sheep, dog and humans. The serodiagnosis of brucellosis involves detection of antibodies generated against the LPS or whole cell bacterial extracts, however these tests lack sensitivity and specificity. The present study was performed to optimize the culture condition for the production of recombinant Brucella melitensis outer membrane protein 28 kDa protein in E.coli via fed batch fermentation. Expression was induced with 1.5mM isopropyl β thiogalactoside and the expressed recombinant protein was purified using Ni-NTA affinity chromatography. After fed-batch fermentation the dry cell weight of 17.81 g/L and a purified protein yield of 210.10 mg/L was obtained. The purified Brucella melitensis recombinant Omp 28 kDa protein was analyzed through SDS- poly acrylamide gel electrophoresis and western blotting. The obtained recombinant protein was evaluated for its diagnostic application through Indirect ELISA using brucellosis suspected human sera samples. Our results clearly indicate that recombinant Omp28 produced via fed batch fermentation has immense potential as a diagnostic reagent that could be employed in sero monitoring of brucellosis.
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B.S. Karothia, T.N. Athmaram, D. Thavaselvam, Ashu Kumar, Tiwari Sapna, K. Singh Anil, K. Sathyaseelan and N. Gopalan, Fed Batch Fermentation and Purification Strategy for High Yield Production of Brucella melitensis Recombinant Omp 28 kDa Protein and its Application in Disease Diagnosis, Protein & Peptide Letters 2013; 20 (7) . https://dx.doi.org/10.2174/0929866511320070001
DOI https://dx.doi.org/10.2174/0929866511320070001 |
Print ISSN 0929-8665 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5305 |
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