EXPRESSION, PURFICATION AND ENDOTOXIN REMOVAL OF BRUCELLA MELITENSIS DNAK AND OMP31 PROTEINS

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GHASEMI A.; SALARI M.H.; ZARNANI A.H.; JEDDI TEHRANI M.

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Journal: Iranian South Medical Journal Year:2014 | Volume:17 | Issue:4 Page(s): 516-523

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Title

EXPRESSION, PURFICATION AND ENDOTOXIN REMOVAL OF BRUCELLA MELITENSIS DNAK AND OMP31 PROTEINS

Author(s)

GHASEMI A. | SALARI M.H. | ZARNANI A.H. | JEDDI TEHRANI M. | Issue Writer Certificate

Keywords

RECOMBINANT PROTEINQ2

PURIFICATIONQ3

ENDOTOXINQ2

EXPRESSIONQ2

Abstract

Background: New strategies are needed to protect against Brucella melitensis infection. Subunit vaccines offer a promising approach because they can stimulate both cellular and humoral immunity, so high production of RECOMBINANT PROTEIN with less content of ENDOTOXIN is desired. In present study, we described a method for EXPRESSION and PURIFICATION of B.melitensis recombinant DnaK(rDnaK) and Omp31(rOmp31) proteins while less content of ENDOTOXINs were detected in final product.Material and Methods: Recombinant pET-dnak and pDEST-omp31 plasmids were transformed into competent EXPRESSION host E.coli BL21 (DE3). After induction by IPTG, bacteria were grown at 20◦C for 22h. Then RECOMBINANT PROTEINs were purified by Ni-NTA Agarose. PURIFICATION was done while two methods using Triton X-114 in washing steps and standard protocol (without detergent) were used in parallel.Results: rDnak and rOmp31 were purified by using Urea. We could obtain 20 and 8 mg RECOMBINANT PROTEINs from rDnak and rOmp31 from 1 liter medium, respectively. The amount of ENDOTOXINs in final products was less content of 0.05 EU/mg. Furthermore, recovery of protein was up to 80% as compared to the standard protocol.Conclusion: The method used in this study, gives a product with very low extent of ENDOTOXIN, but 20% of RECOMBINANT PROTEINs were lost. So we think the method described here can be used for PURIFICATION and ENDOTOXIN removal of other RECOMBINANT PROTEINs with similar physiologic properties.

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APA: Copy

GHASEMI, A., SALARI, M.H., ZARNANI, A.H., & JEDDI TEHRANI, M.. (2014). EXPRESSION, PURFICATION AND ENDOTOXIN REMOVAL OF BRUCELLA MELITENSIS DNAK AND OMP31 PROTEINS. IRANIAN SOUTH MEDICAL JOURNAL (ISMJ), 17(4), 516-523. SID. https://sid.ir/paper/33499/en

Vancouver: Copy

GHASEMI A., SALARI M.H., ZARNANI A.H., JEDDI TEHRANI M.. EXPRESSION, PURFICATION AND ENDOTOXIN REMOVAL OF BRUCELLA MELITENSIS DNAK AND OMP31 PROTEINS. IRANIAN SOUTH MEDICAL JOURNAL (ISMJ)[Internet]. 2014;17(4):516-523. Available from: https://sid.ir/paper/33499/en

IEEE: Copy

A. GHASEMI, M.H. SALARI, A.H. ZARNANI, and M. JEDDI TEHRANI, “EXPRESSION, PURFICATION AND ENDOTOXIN REMOVAL OF BRUCELLA MELITENSIS DNAK AND OMP31 PROTEINS,” IRANIAN SOUTH MEDICAL JOURNAL (ISMJ), vol. 17, no. 4, pp. 516–523, 2014, [Online]. Available: https://sid.ir/paper/33499/en

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