Micropreparative fractionation of the complexome by blue native continuous elution electrophoresis

Huang, KY; Filarsky, M; Padula, MP; Raftery, MJ; Herbert, BR; Wilkins, MR

Micropreparative fractionation of the complexome by blue native continuous elution electrophoresis

Huang, KY Filarsky, M Padula, MP

Raftery, MJ Herbert, BR

Wilkins, MR

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Publication Type:: Journal Article
Citation:: Proteomics, 2009, 9 (9), pp. 2494 - 2502
Issue Date:: 2009-05-01

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Field	Value	Language
dc.contributor.author	Huang, KY	en_US
dc.contributor.author	Filarsky, M	en_US
dc.contributor.author	Padula, MP https://orcid.org/0000-0002-8283-0643	en_US
dc.contributor.author	Raftery, MJ	en_US
dc.contributor.author	Herbert, BR https://orcid.org/0000-0003-4697-8197	en_US
dc.contributor.author	Wilkins, MR	en_US
dc.date.issued	2009-05-01	en_US
dc.identifier.citation	Proteomics, 2009, 9 (9), pp. 2494 - 2502	en_US
dc.identifier.issn	1615-9853	en_US
dc.identifier.uri	http://hdl.handle.net/10453/29880
dc.description.abstract	The large-scale analysis of protein complexes is an emerging challenge in the field of proteomics. Currently, there are few methods available for the fractionation of protein complexes that are compatible with downstream proteomic techniques. Here, we describe the technique of blue native continuous elution electrophoresis (BN-CEE). It combines the features of blue native PAGE (BN-PAGE) and continuous elution electrophoresis (CEE), generating liquid-phase fractions of protein complexes of up to 800 kDa. The resulting complexes can be further analysed by BN-PAGE, by SDS-PAGE and/or by MS. This can help define the constituent proteins of many complexes and their stoichiometry. As BN-CEE is also micropreparative, with a capacity to separate milligram quantities ofprotein complexes, it will assist the study ofproteins oflower abundance. In this regard, the acrylamide concentration and elution rate during separation can be controlled to help 'zoom in' on particular high mass regions and thus complexes of interest. We illustrate the utility of the technique in the analysis of Saccharomyces cerevisiae cellular lysate. © 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.	en_US
dc.relation.ispartof	Proteomics	en_US
dc.relation.isbasedon	10.1002/pmic.200800525	en_US
dc.subject.classification	Biochemistry & Molecular Biology	en_US
dc.subject.mesh	Multiprotein Complexes	en_US
dc.subject.mesh	Proteins	en_US
dc.subject.mesh	Saccharomyces cerevisiae Proteins	en_US
dc.subject.mesh	Chemical Fractionation	en_US
dc.subject.mesh	Electrophoresis, Polyacrylamide Gel	en_US
dc.subject.mesh	Proteomics	en_US
dc.subject.mesh	Molecular Weight	en_US
dc.subject.mesh	Mass Spectrometry	en_US
dc.title	Micropreparative fractionation of the complexome by blue native continuous elution electrophoresis	en_US
dc.type	Journal Article
utslib.citation.volume	9	en_US
utslib.for	0601 Biochemistry and Cell Biology	en_US
utslib.for	06 Biological Sciences	en_US
utslib.for	08 Information and Computing Sciences	en_US
utslib.for	11 Medical and Health Sciences	en_US
pubs.embargo.period	Not known	en_US
pubs.organisational-group	/University of Technology Sydney
pubs.organisational-group	/University of Technology Sydney/Faculty of Science
pubs.organisational-group	/University of Technology Sydney/Faculty of Science/School of Life Sciences
utslib.copyright.status	closed_access
pubs.issue	9	en_US
pubs.publication-status	Published	en_US
pubs.volume	9	en_US

Abstract:

The large-scale analysis of protein complexes is an emerging challenge in the field of proteomics. Currently, there are few methods available for the fractionation of protein complexes that are compatible with downstream proteomic techniques. Here, we describe the technique of blue native continuous elution electrophoresis (BN-CEE). It combines the features of blue native PAGE (BN-PAGE) and continuous elution electrophoresis (CEE), generating liquid-phase fractions of protein complexes of up to 800 kDa. The resulting complexes can be further analysed by BN-PAGE, by SDS-PAGE and/or by MS. This can help define the constituent proteins of many complexes and their stoichiometry. As BN-CEE is also micropreparative, with a capacity to separate milligram quantities ofprotein complexes, it will assist the study ofproteins oflower abundance. In this regard, the acrylamide concentration and elution rate during separation can be controlled to help 'zoom in' on particular high mass regions and thus complexes of interest. We illustrate the utility of the technique in the analysis of Saccharomyces cerevisiae cellular lysate. © 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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http://hdl.handle.net/10453/29880