Optimization of Human Interferon α2b Soluble Protein Overproduction and Primary Recovery of Its Inclusion Bodies

Authors

  • RATIH ASMANA NINGRUM School of Pharmacy, Institut Teknologi Bandung
  • DEBBIE SOFIE RETNONINGRUM School of Pharmacy, Institut Teknologi Bandung
  • YEYET CAHYATI School of Pharmacy, Institut Teknologi Bandung
  • HENI RACHMAWATI School of Pharmacy, Institut Teknologi Bandung

DOI:

https://doi.org/10.5454/mi.5.1.5

Keywords:

human interferon α2b, overproduction, soluble protein, inclusion bodies, protein refolding

Abstract

The hIFN2b open reading frame has been constructed and overexpressed in Escherichia coli BL21(DE3). The yields of protein purified using nickel column from inclusion bodies (IB) and total soluble proteins were 3.46 mg and 2.57 mg in 1 L culture, respectively. This research was aimed to obtain optimal condition for high level overproduction of soluble hIFNα2b as well as primary recovery of hIFN2b from IB. We used two different conditions for obtaining soluble protein, i.e. induction temperatures and inducer concentrations, and three different conditions for inclusion bodies, i.e. centrifugation speeds, washing and solubilizing buffers. Induction using 0.5 mM of isopropyl thiogalactopyranoside at 25 °C yielded 8.9 mg hIFN2b in 1 L culture. The best recovery of IB was achieved when 10 000 g was applied for centrifugation, 1% Triton X-100  in 50 mM Tris Cl pH 8.0 as washing buffer, and 8M guanidine HCl in 50 mM Tris Cl pH 8.0 containing 800 mM 2-mercaptoethanol as solubilizing buffer were used. At this optimal condition the yield of hIFN2b from IB was 28.85 mg in 1 L culture. The total recovery of hIFNα2b at optimal condition was 50% from IB and 14% from soluble protein. hIFN2b from IB was refolded by 9 d dialysis in refolding buffer (0.2 mM EDTA, 0.25 mM ditiothreitol, 50 mM Tris and 0.4 M urea pH 8.0).

Downloads

Published

2011-04-25

How to Cite

NINGRUM, R. A., RETNONINGRUM, D. S., CAHYATI, Y., & RACHMAWATI, H. (2011). Optimization of Human Interferon α2b Soluble Protein Overproduction and Primary Recovery of Its Inclusion Bodies. Microbiology Indonesia, 5(1), 5. https://doi.org/10.5454/mi.5.1.5

Issue

Section

Articles
                                                                  ADOConnection.CacheExecute(3600, SELECT 				o.submission_id, 				MAX(s.context_id) AS journal_id, 				MAX(i.date_published) AS i_pub, 				MAX(p.date_published) A..., Array[2]) % line  132, file: DAO.inc.php
                                                               DAO.retrieveCached(SELECT 				o.submission_id, 				MAX(s.context_id) AS journal_id, 				MAX(i.date_published) AS i_pub, 				MAX(p.date_published) A..., Array[2], 3600) % line   91, file: ArticleSearchDAO.inc.php
                                                            ArticleSearchDAO.getPhraseResults(Object:Journal, Array[1], null, null, , 500, 1) % line  201, file: SubmissionSearch.inc.php
                                                         SubmissionSearch._getMergedPhraseResults(Object:Journal, Array[1], , null, null, 500, 1) % line  146, file: SubmissionSearch.inc.php
                                                      SubmissionSearch._getMergedKeywordResults(Object:Journal, Array[4], , null, null, 500, 1) % line  194, file: SubmissionSearch.inc.php
                                                   SubmissionSearch._getMergedPhraseResults(Object:Journal, Array[4], null, null, null, 500, 1) % line  146, file: SubmissionSearch.inc.php
                                                SubmissionSearch._getMergedKeywordResults(Object:Journal, Array[3], null, null, null, 500, 1) % line  132, file: SubmissionSearch.inc.php
                                             SubmissionSearch._getMergedArray(Object:Journal, Array[1], null, null) % line  263, file: SubmissionSearch.inc.php
                                          SubmissionSearch.retrieveResults(Object:Request, Object:Journal, Array[1], null, null, null, Object:DBResultRange, Array[1]) % line   92, file: RecommendBySimilarityPlugin.inc.php
                                       RecommendBySimilarityPlugin.callbackTemplateArticlePageFooter(Templates::Article::Footer::PageFooter, Array[3]) % line  107, file: HookRegistry.inc.php
                                    HookRegistry.call(Templates::Article::Footer::PageFooter, Array[3]) % line 1256, file: PKPTemplateManager.inc.php
                                 PKPTemplateManager.smartyCallHook(Array[1], Object:Smarty_Internal_Template) % line   43, file: 42362804174f266c598cdb32c3208fc745b92101^1c372c95cd85572e0fbc9a53d0323a0b229cfc78_0.app.frontendpagesarticle.tpl.php
                              content_6082325297a527_48332634(Object:Smarty_Internal_Template) % line  123, file: smarty_template_resource_base.php
                           Smarty_Template_Resource_Base.getRenderedTemplateCode(Object:Smarty_Internal_Template) % line  114, file: smarty_template_compiled.php
                        Smarty_Template_Compiled.render(Object:Smarty_Internal_Template) % line  216, file: smarty_internal_template.php
                     Smarty_Internal_Template.render(false, 1) % line  232, file: smarty_internal_templatebase.php
                  Smarty_Internal_TemplateBase._execute(Object:Smarty_Internal_Template, null, 42362804174f266c598cdb32c3208fc745b92101, null, 1) % line  134, file: smarty_internal_templatebase.php
               Smarty_Internal_TemplateBase.display(frontend/pages/article.tpl, null, 42362804174f266c598cdb32c3208fc745b92101, null) % line  924, file: PKPTemplateManager.inc.php
            PKPTemplateManager.display(frontend/pages/article.tpl) % line  287, file: ArticleHandler.inc.php
         ArticleHandler.view(Array[1], Object:Request) % line  391, file: PKPRouter.inc.php
      PKPRouter._authorizeInitializeAndCallRequest(Array[2], Object:Request, Array[1], false) % line  231, file: PKPPageRouter.inc.php
   PKPPageRouter.route(Object:Request) % line  143, file: Dispatcher.inc.php
Dispatcher.dispatch(Object:Request) % line  281, file: PKPApplication.inc.php
PKPApplication.execute() % line   68, file: index.php

Similar Articles

You may also start an advanced similarity search for this article.