Solubilisation of the armadillo‐repeat protein β‐catenin using a zwitterionic detergent allows resolution of phosphorylated forms by 2DE

Electrophoresis - Tập 33 Số 12 - Trang 1804-1813 - 2012
Meredith J. Layton1, Nicole Church1, Maree C. Faux1, Hong Ji2,1, Robert J. A. Goode1, Eugene A. Kapp1, Antony W. Burgess1, Richard J. Simpson2,1
1The Ludwig Institute for Cancer Research, Royal Melbourne Hospital, Parkville, Australia.
2La Trobe Institute for Molecular Science (LIMS) La Trobe University Bundoora Australia

Tóm tắt

β‐catenin is a member of the armadillo repeat family of proteins and has important functions in cell–cell adhesion and Wnt signalling. Different protein species of β‐catenin have been shown to exist in the cell and the relative proportions of these species are altered upon stimulation of cells with Wnt‐3a (Gottardi and Gumbiner, 2004). In order to determine whether posttranslational modifications (PTMs) of β‐catenin underlie these different protein species, we have used 2DE separation and immunoblotting with an antibody specific for β‐catenin. High‐resolution separation of differentially modified species of β‐catenin in 2DE required the addition of ASB‐16, a zwitterionic detergent that can solubilise integral membrane proteins. ASB‐16 was also necessary for focussing of other armadillo repeat proteins, such as γ‐catenin and p120‐catenin. 2DE using ASB‐16 allowed detection of a previously unreported phosphorylation site in the transcriptionally active form of β‐catenin that binds to GST‐Tcf in response to Wnt signalling.

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Electrophoresis

Tập 33 Số 12

1804-1813

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