Autor(es): Tripathi Nagesh K, Priya Raj, Shrivastava Ambuj

Resumo: Chikungunya, a mosquito-borne viral disease caused by Chikungunya virus (CHIKV), has drawn substantial attention after its reemergence causing massive outbreaks in tropical regions of Asia - Africa. The recombinant envelope 2 (rE2) protein of CHIKV is a potential diagnostic as well as vaccine c-idate. Development of cost-effective cultivation media - appropriate culture conditions are generally favorable for large-scale production of recombinant proteins in Escherichia coli. The effects of medium composition - cultivation conditions on the production of recombinant Chikungunya virus E2 (rCHIKV E2) protein were investigated in shake flask culture as well as batch cultivation of Escherichia coli. Further, the fed-batch process was also carried out for high cell density cultivation of E. coli expressing rE2 protein. Expression of rCHIKV E2 protein in E. coli was induced with 1 mM isopropyl-beta-thiogalactoside (IPTG) at ~23 g dry cell weight (DCW) per liter of culture - yielded an insoluble protein aggregating to form inclusion bodies. The final DCW after fed-batch cultivation was ~35 g/l. The inclusion bodies were isolated, solubilized in 8 M urea - purified through affinity chromatography to give a final product yield of ~190 mg/l. The reactivity of purified E2 protein was confirmed by Western blotting - enzyme-linked immunosorbent assay. These results show that rE2 protein of CHIKV may be used as a diagnostic reagent or for further prophylactic studies. This approach of producing rE2 protein in E. coli with high yield may also offer a promising method for production of other viral recombinant proteins.

Palavras-Chave: Chikungunya virus escherichia coli bioreactor cultivation ELISA

Imprenta: Applied Microbiology and Biotechnology, v. 98, n. 6, p. 2461-2471, 2014

Identificador do Objeto Digital: 10.1007/s00253-013-5426-4

Descritores: Chikungunya virus - Biosynthesis ; Chikungunya virus - Cell ; Chikungunya virus - Pathogenesis ; Chikungunya virus - Proteins ; Chikungunya Virus - Virus ; Chikungunya virus - Vaccine

Data de Publicação: 2014