Direct refolding of inclusion bodies using reversed micelles

Masafumi Sakono, Yu Mi Kawashima, Hirofumi Ichinose, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

The protein refolding of inclusion bodies was investigated using reversed micelles formed by aerosol OT (AOT). Ribonuclease A (RNase A) was overexpressed in Escherichia coli and used as native inclusion bodies. The enzymatic activity of RNase A was completely regained from the inclusion bodies within 14 h by solubilization in reversed micelles. To further enhance the refolding rate, a molecular chaperone, GroEL, was incorporated into the refolding system. The resultant refolding system including GroEL showed better performance under optimized conditions for the refolding of RNase A inclusion bodies. The refolding rate was considerably improved by the addition of the molecular chaperone, and the refolding step was completed in 1 h. The protein refolding in the GroEL-containing refolding system was strongly dependent on the coexistence of ATP and Mg2+, suggesting that the GroEL hosted in the reversed micelles was biologically active and assisted in the renaturation of the inclusion bodies. The addition of cold acetone to the reversed micellar solution allowed over 90% recovery of the renatured RNase A.

Original languageEnglish
Pages (from-to)1783-1787
Number of pages5
JournalBiotechnology Progress
Volume20
Issue number6
DOIs
Publication statusPublished - Nov 1 2004

All Science Journal Classification (ASJC) codes

  • Biotechnology

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