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Forler, D., Koecher, T., Rode, M., Gentzel, M., Izaurralde, E., Wilm, M. (2003). An efficient protein complex purification method for functional proteomics in higher eukaryotes.  Nat. Biotechnol. 21(1): 89--92.
FlyBase ID
FBrf0155992
Publication Type
Research paper
Abstract

The ensemble of expressed proteins in a given cell is organized in multiprotein complexes. The identification of the individual components of these complexes is essential for their functional characterization. The introduction of the 'tandem affinity purification' (TAP) methodology substantially improved the purification and systematic genome-wide characterization of protein complexes in yeast. The use of this approach in higher eukaryotic cells has lagged behind its use in yeast because the tagged proteins are normally expressed in the presence of the untagged endogenous version, which may compete for incorporation into multiprotein complexes. Here we describe a strategy in which the TAP approach is combined with double-stranded RNA interference (RNAi) to avoid competition from corresponding endogenous proteins while isolating and characterizing protein complexes from higher eukaryotic cells. This strategy allows the determination of the functionality of the tagged protein and increases the specificity and the efficiency of the purification.

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    Language of Publication
    English
    Additional Languages of Abstract
    Parent Publication
    Publication Type
    Journal
    Abbreviation
    Nat. Biotechnol.
    Title
    Nature biotechnology
    Publication Year
    1996-
    ISBN/ISSN
    1087-0156
    Data From Reference
    Gene Groups (1)
    Genes (13)