Calixarene-assisted protein refolding via liquid-liquid extraction.

Abstract:

:In this paper we report on protein refolding by means of a liquid-liquid transfer technique using a calixarene. We have found that a calix[6]areneacetic acid derivative forms a supramolecular complex with urea-denatured cytochrome c at the oil-water interface, which enables quantitative transfer of the protein from an 8 M urea aqueous solution into an organic phase through a proton-exchange mechanism. Denatured cytochrome c is completely separated from the denaturant and is isolated from other denatured cytochrome c molecules to suppress the generation of aggregates due to protein-protein interactions. The recovery of cytochrome c from the organic phase is successfully achieved under acidic conditions using an appropriate amount of 1-butanol. UV-vis, CD, and fluorescence spectroscopic characterizations demonstrate that cytochrome c transferred into a denaturant-free aqueous solution regains its native structure. The reduction kinetics of refolded cytochrome c using ascorbic acid indicates that the protein provides approximately 72% of native activity as an electron-transfer protein.

journal_name

Biomacromolecules

journal_title

Biomacromolecules

authors

Shimojo K,Oshima T,Naganawa H,Goto M

doi

10.1021/bm070418q

subject

Has Abstract

pub_date

2007-10-01 00:00:00

pages

3061-6

issue

10

eissn

1525-7797

issn

1526-4602

journal_volume

8

pub_type

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