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The Protein Journal

, Volume 29, Issue 7, pp 457–465 | Cite as

Mechanism of Gemini Disulfide Detergent Mediated Oxidative Refolding of Lysozyme in a New Artificial Chaperone System

  • Marc Potempa
  • Mathias Hafner
  • Christian Frech
Article

Abstract

Gemini surfactants are a new class of surfactants that consist of two hydrophilic head groups and two hydrophobic tails separated by a spacer group. As the properties of geminis are different to their monomeric counterparts, a large number of applications have been investigated. Here we report on the use of a new class of gemini detergents containing a disulfide bond in the spacer (Det-SS-Det) for protein refolding. Using lysozyme as a model protein we could demonstrate that the disulfide gemini detergents allow oxidative refolding of the protein in the absence of any external redox system in an “artificial chaperone system”. Refolding kinetics using gemini disulfide detergents differing in their hydrophobicity were analysed to determine the folding and aggregation rate constants. The results point to an important role of the transiently formed mixed disulfides between the protein and the detergent (Prot-SS-Det) in the oxidative refolding process of lysozyme.

Keywords

Disulfide gemini detergent Artificial chaperone Oxidative refolding Lysozyme 

Abbreviations

ACN

Acetonitrile

ALP

Alkaline phosphatase

β-CD

β-cyclodextrin

BSA

Bovine serum albumin

CD

Cyclodextrin

CMC

Critical micelle concentration

CTAB

Cetyltrimethylammonium bromide

C9SS

Nonyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C10SS

Decyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C11SS

Undecyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C12SS

Dodecyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C13SS

Tridecyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C14SS

Tetradecyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C15SS

Pentadecyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

C16SS

Hexadecyl-dimethyl-(2-sulfanylethyl)azanium-disulfide

DTAB

Dodecyltrimethylammonium bromide

DTTox

Oxidized dithiothreitol

DTTred

Reduced dithiothreitol

E. coli

Escherichia coli

EDTA

Ethylenediaminetetraacetic acid

GdmCl

Guanidine hydrochloride

GSH

Reduced glutathione

GSSG

Oxidized glutathione

HPLC

High pressure liquid chromatography

IB

Inclusion body

m-β-CD

Methyl-β-cyclodextrin

RNAse A

Ribonuclease A

RP-HPLC

Reversed phase high pressure liquid chromatography

RT

Room temperature

TCEP

Tris (2-carboxyethyl) phosphine hydrochloride

TFA

Trifluoroacetic acid

Tris

Tris (hydroxymethyl)-aminomethan

TTAB

Tetradecyltrimethylammonium bromide

Notes

Acknowledgments

This work was supported by the Landesstiftung Baden-Württemberg.

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Institute for BiochemistryUniversity of Applied SciencesMannheimGermany
  2. 2.Institute of Molecular Biology and Cell Culture TechnologyUniversity of Applied SciencesMannheimGermany

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