Advanced search
Start date
(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Protein refolding based on high hydrostatic pressure and alkaline pH: Application on a recombinant dengue virus NS1 protein

Full text
Chura-Chambi, Rosa Maria [1] ; Rosa da Silva, Cleide Mara [1] ; Pereira, Lennon Ramos [2] ; Bartolini, Paolo [1] ; de Souza Ferreira, Luis Carlos [2] ; Morganti, Ligia [1]
Total Authors: 6
[1] IPEN CNEN SP, Inst Pesquisas Energet & Nucl, Ctr Biotecnol, Sao Paulo, SP - Brazil
[2] Univ Sao Paulo, Inst Ciencias Biomed, Dept Microbiol, Sao Paulo, SP - Brazil
Total Affiliations: 2
Document type: Journal article
Source: PLoS One; v. 14, n. 1 JAN 25 2019.
Web of Science Citations: 3

In this study we evaluated the association of high hydrostatic pressure (HHP) and alkaline pH as a minimally denaturing condition for the solubilization of inclusion bodies (IBs) generated by recombinant proteins expressed by Escherichia coli strains. The method was successfully applied to a recombinant form of the dengue virus (DENV) non-structural protein 1 (NS1). The minimal pH for IBs solubilization at 1 bar was 12 while a pH of 10 was sufficient for solubilization at HHP: 2.4 kbar for 90 min and 0.4 kbar for 14 h 30 min. An optimal refolding condition was achieved by compression of IBs at HHP and pH 10.5 in the presence of arginine, oxidized and reduced glutathiones, providing much higher yields (up to 8-fold) than association of HHP and GdnHCl via an established protocol. The refolded NS1, 109 +/- 9.5 mg/L bacterial culture was recovered mainly as monomer and dimer, corresponding up to 90% of the total protein and remaining immunologically active. The proposed conditions represent an alternative for the refolding of immunologically active recombinant proteins expressed as IBs. (AU)

FAPESP's process: 15/02574-0 - Refolding of antigens synthesized as inclusion bodies in Escherichia coli for vaccine preparation
Grantee:Ligia Ely Morganti Ferreira Dias
Support Opportunities: Regular Research Grants