title: Tris-acetate polyacrylamide gradient gel electrophoresis for the analysis of protein oligomerization.
abstract: Here we report a new approach for studying protein oligomerization in cells using a single electrophoresis gel. We combined the use of a crosslinking reagent for sample preparation, such as glutaraldehyde, with the analysis of oligomers by Tris-acetate polyacrylamide gel electrophoresis. The use of a 3-15% Tris-acetate polyacrylamide gradient gel allows for the simultaneous analysis of proteins of masses ranging from 10 to 500 kDa. We showed the usefulness of this method for analyzing endogenous p53 oligomerization with high resolution and sensitivity in human cells. Oligomerization analysis was dependent on the crosslinker concentration used. We also showed that this method could be used to study the regulation of oligomerization. In all experiments, Tris-acetate polyacrylamide gel electrophoresis proved to be a robust, manageable, and cost- and time-efficient method that provided excellent results using a single gel. This approach can be easily extrapolated to the study of other oligomers. All of these features make this method a highly useful tool for the analysis of protein oligomerization.
authors.affiliation: Departament de Ciències Fisiològiques II, IDIBELL, Campus de Bellvitge, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Barcelona, Spain.
authors.forename: Monica
authors.initials: M
authors.lastname: Cubillos-Rojas
authors.affiliation: Departament de Ciències Fisiològiques II, IDIBELL, Campus de Bellvitge, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Barcelona, Spain.
authors.forename: Taiane
authors.initials: T
authors.lastname: Schneider
authors.affiliation: Departament de Ciències Fisiològiques II, IDIBELL, Campus de Bellvitge, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Barcelona, Spain.
authors.forename: Susana
authors.initials: S
authors.lastname: Sánchez-Tena
authors.affiliation: Departament de Ciències Fisiològiques II, IDIBELL, Campus de Bellvitge, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Barcelona, Spain.
authors.forename: Ramon
authors.initials: R
authors.lastname: Bartrons
authors.affiliation: Departament de Ciències Fisiològiques II, IDIBELL, Campus de Bellvitge, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Barcelona, Spain.
authors.forename: Francesc
authors.initials: F
authors.lastname: Ventura
authors.affiliation: Departament de Ciències Fisiològiques II, IDIBELL, Campus de Bellvitge, Universitat de Barcelona, 08907 L'Hospitalet de Llobregat, Barcelona, Spain. joseluisrosa@ub.edu.
authors.forename: Jose Luis
authors.initials: JL
authors.lastname: Rosa
mesh_terms: D000085:Acetates / Q000737:chemistry; D001761:Bleomycin / Q000494:pharmacology; D002460:Cell Line; D003432:Cross-Linking Reagents / Q000737:chemistry; D004317:Doxorubicin / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005976:Glutaral / Q000737:chemistry; D006801:Humans; D008970:Molecular Weight; D055503:Protein Multimerization / Q000187:drug effects; D011506:Proteins / Q000032:analysis* / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000032:analysis / Q000378:metabolism
chemical_list: D000085:Acetates; D003432:Cross-Linking Reagents; D011506:Proteins; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D001761:Bleomycin; D004317:Doxorubicin; D005976:Glutaral
keywords: Gradient gel; Oligomerization; PAGE; Protein crosslinking; Tris-acetate