Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus
Abstract Background Proteins in inclusion bodies (IBs) present native-like secondary structures. However, chaotropic agents at denaturing concentrations, which are widely used for IB solubilization and subsequent refolding, unfold these secondary structures. Removal of the chaotropes frequently caus...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2018-12-01
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| Series: | BMC Biotechnology |
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| Online Access: | http://link.springer.com/article/10.1186/s12896-018-0486-2 |
| _version_ | 1818878859564548096 |
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| author | Cleide Mara Rosa da Silva Rosa Maria Chura-Chambi Lennon Ramos Pereira Yraima Cordeiro Luís Carlos de Souza Ferreira Ligia Morganti |
| author_facet | Cleide Mara Rosa da Silva Rosa Maria Chura-Chambi Lennon Ramos Pereira Yraima Cordeiro Luís Carlos de Souza Ferreira Ligia Morganti |
| author_sort | Cleide Mara Rosa da Silva |
| collection | DOAJ |
| description | Abstract Background Proteins in inclusion bodies (IBs) present native-like secondary structures. However, chaotropic agents at denaturing concentrations, which are widely used for IB solubilization and subsequent refolding, unfold these secondary structures. Removal of the chaotropes frequently causes reaggregation and poor recovery of bioactive proteins. High hydrostatic pressure (HHP) and alkaline pH are two conditions that, in the presence of low level of chaotropes, have been described as non-denaturing solubilization agents. In the present study we evaluated the strategy of combination of HHP and alkaline pH on the solubilization of IB using as a model an antigenic form of the zika virus (ZIKV) non-structural 1 (NS1) protein. Results Pressure-treatment (2.4 kbar) of NS1-IBs at a pH of 11.0 induced a low degree of NS1 unfolding and led to solubilization of the IBs, mainly into monomers. After dialysis at pH 8.5, NS1 was refolded and formed soluble oligomers. High (up to 68 mg/liter) NS1 concentrations were obtained by solubilization of NS1-IBs at pH 11 in the presence of arginine (Arg) with a final yield of approximately 80% of total protein content. The process proved to be efficient, quick and did not require further purification steps. Refolded NS1 preserved biological features regarding reactivity with antigen-specific antibodies, including sera of ZIKV-infected patients. The method resulted in an increase of approximately 30-fold over conventional IB solubilization-refolding methods. Conclusions The present results represent an innovative non-denaturing protein refolding process by means of the concomitant use of HHP and alkaline pH. Application of the reported method allowed the recovery of ZIKV NS1 at a condition that maintained the antigenic properties of the protein. |
| first_indexed | 2024-12-19T14:20:53Z |
| format | Article |
| id | doaj.art-80bef607c3644fd2b2df75b027b9ae1d |
| institution | Directory Open Access Journal |
| issn | 1472-6750 |
| language | English |
| last_indexed | 2024-12-19T14:20:53Z |
| publishDate | 2018-12-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Biotechnology |
| spelling | doaj.art-80bef607c3644fd2b2df75b027b9ae1d2022-12-21T20:17:46ZengBMCBMC Biotechnology1472-67502018-12-0118111010.1186/s12896-018-0486-2Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virusCleide Mara Rosa da Silva0Rosa Maria Chura-Chambi1Lennon Ramos Pereira2Yraima Cordeiro3Luís Carlos de Souza Ferreira4Ligia Morganti5Instituto de Pesquisas Energéticas e Nucleares, IPEN-CNEN/SP, Centro de BiotecnologiaInstituto de Pesquisas Energéticas e Nucleares, IPEN-CNEN/SP, Centro de BiotecnologiaDepartamento de Microbiologia, Universidade de São Paulo, Instituto de Ciências BiomédicasUniversidade Federal do Rio de Janeiro, Faculdade de FarmáciaDepartamento de Microbiologia, Universidade de São Paulo, Instituto de Ciências BiomédicasInstituto de Pesquisas Energéticas e Nucleares, IPEN-CNEN/SP, Centro de BiotecnologiaAbstract Background Proteins in inclusion bodies (IBs) present native-like secondary structures. However, chaotropic agents at denaturing concentrations, which are widely used for IB solubilization and subsequent refolding, unfold these secondary structures. Removal of the chaotropes frequently causes reaggregation and poor recovery of bioactive proteins. High hydrostatic pressure (HHP) and alkaline pH are two conditions that, in the presence of low level of chaotropes, have been described as non-denaturing solubilization agents. In the present study we evaluated the strategy of combination of HHP and alkaline pH on the solubilization of IB using as a model an antigenic form of the zika virus (ZIKV) non-structural 1 (NS1) protein. Results Pressure-treatment (2.4 kbar) of NS1-IBs at a pH of 11.0 induced a low degree of NS1 unfolding and led to solubilization of the IBs, mainly into monomers. After dialysis at pH 8.5, NS1 was refolded and formed soluble oligomers. High (up to 68 mg/liter) NS1 concentrations were obtained by solubilization of NS1-IBs at pH 11 in the presence of arginine (Arg) with a final yield of approximately 80% of total protein content. The process proved to be efficient, quick and did not require further purification steps. Refolded NS1 preserved biological features regarding reactivity with antigen-specific antibodies, including sera of ZIKV-infected patients. The method resulted in an increase of approximately 30-fold over conventional IB solubilization-refolding methods. Conclusions The present results represent an innovative non-denaturing protein refolding process by means of the concomitant use of HHP and alkaline pH. Application of the reported method allowed the recovery of ZIKV NS1 at a condition that maintained the antigenic properties of the protein.http://link.springer.com/article/10.1186/s12896-018-0486-2Protein refoldingInclusion bodiesNS1Dengue virus and Zika virusHigh hydrostatic pressureAlkaline pH |
| spellingShingle | Cleide Mara Rosa da Silva Rosa Maria Chura-Chambi Lennon Ramos Pereira Yraima Cordeiro Luís Carlos de Souza Ferreira Ligia Morganti Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus BMC Biotechnology Protein refolding Inclusion bodies NS1 Dengue virus and Zika virus High hydrostatic pressure Alkaline pH |
| title | Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus |
| title_full | Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus |
| title_fullStr | Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus |
| title_full_unstemmed | Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus |
| title_short | Association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the NS1 protein from zika virus |
| title_sort | association of high pressure and alkaline condition for solubilization of inclusion bodies and refolding of the ns1 protein from zika virus |
| topic | Protein refolding Inclusion bodies NS1 Dengue virus and Zika virus High hydrostatic pressure Alkaline pH |
| url | http://link.springer.com/article/10.1186/s12896-018-0486-2 |
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