Refolding of a membrane protein in a microfluidics reactor

Zaccai, Nathan R., Yunus, Kamran, Matthews, S. M., Fisher, Adrian C. and Falconer, Robert J. (2007) Refolding of a membrane protein in a microfluidics reactor. European Biophysics Journal With Biophysics Letters, 36 6: 581-588. doi:10.1007/s00249-006-0125-z


Author Zaccai, Nathan R.
Yunus, Kamran
Matthews, S. M.
Fisher, Adrian C.
Falconer, Robert J.
Title Refolding of a membrane protein in a microfluidics reactor
Journal name European Biophysics Journal With Biophysics Letters   Check publisher's open access policy
ISSN 0175-7571
Publication date 2007-07-01
Sub-type Article (original research)
DOI 10.1007/s00249-006-0125-z
Open Access Status DOI
Volume 36
Issue 6
Start page 581
End page 588
Total pages 8
Place of publication New York
Publisher Springer
Language eng
Subject 100302 Bioprocessing, Bioproduction and Bioproducts
Abstract Membrane protein production for structural studies is often hindered by the formation of nonspecific aggregates from which the protein has to be denatured and then refolded to a functional state. We developed a new approach, which uses microfluidics channels, to refold protein correctly in quantities sufficient for structural studies. Green fluorescent protein (GFP), a soluble protein, and bacteriorhodopsin (BR), a transmembrane protein, were used to demonstrate the efficiency of the process. Urea-denatured GFP refolded as the urea diffused away from the protein, forming in the channel a uniform fluorescent band when observed by confocal microscopy. Sodium dodecyl sulphate-denatured BR refolded within the channel on mixing with detergent-lipid mixed micelles. The refolding, monitored by absorbance spectroscopy, was found to be flow rate dependent. This potential of microfluidic reactors for screening protein folding conditions and producing protein would he particularly amenable for high-throughput applications required in structural genomics.
Keyword Biophysics
green fluorescent protein
bacteriorhodopsin
microfluidics reactor
membrane
protein refolding
structural genomics
Green-fluorescent Protein
Optical Lithography
Retinal-binding
Folding Screen
In-vitro
Bacteriorhodopsin
Chromophore
Mechanism
Aequorea
Receptor
Q-Index Code C1

Document type: Journal Article
Sub-type: Article (original research)
Collections: Excellence in Research Australia (ERA) - Collection
Australian Institute for Bioengineering and Nanotechnology Publications
 
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Created: Tue, 19 Feb 2008, 00:24:54 EST