Native GELFrEE: A new separation technique for biomolecular assemblies

Owen S. Skinner, Luis H.F. Do Vale, Adam D. Catherman, Pierre C. Havugimana, Marcelo Valle De Sousa, Philip D. Compton*, Neil L. Kelleher

*Corresponding author for this work

Research output: Contribution to journalArticle

21 Scopus citations

Abstract

The cadre of protein complexes in cells performs an array of functions necessary for life. Their varied structures are foundational to their ability to perform biological functions, lending great import to the elucidation of complex composition and dynamics. Native separation techniques that are operative on low sample amounts and provide high resolution are necessary to gain valuable data on endogenous complexes. Here, we detail and optimize the use of tube gel separations to produce samples proven compatible with native, multistage mass spectrometry (nMS/MS). We find that a continuous system (i.e., no stacking gel) with a gradient in its extent of cross-linking and use of the clear native buffer system performs well for both fractionation and native mass spectrometry of heart extracts and a fungal secretome. This integrated advance in separations and nMS/MS offers the prospect of untargeted proteomics at the next hierarchical level of protein organization in biology.

Original languageEnglish (US)
Pages (from-to)3032-3038
Number of pages7
JournalAnalytical Chemistry
Volume87
Issue number5
DOIs
StatePublished - Mar 3 2015

ASJC Scopus subject areas

  • Analytical Chemistry

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    Skinner, O. S., Do Vale, L. H. F., Catherman, A. D., Havugimana, P. C., Sousa, M. V. D., Compton, P. D., & Kelleher, N. L. (2015). Native GELFrEE: A new separation technique for biomolecular assemblies. Analytical Chemistry, 87(5), 3032-3038. https://doi.org/10.1021/ac504678d