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Molecular & Cellular Proteomics 1:204-212, 2002.
© 2002 by The American Society for Biochemistry and Molecular Biology, Inc.

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Research

Deciphering Protein Complexes and Protein Interaction Networks by Tandem Affinity Purification and Mass Spectrometry

Analytical Perspective^*

Anna Shevchenko, Daniel Schaft, Assen Roguev, W. W. M. Pim Pijnappel, A. Francis Stewart^,¶ and Andrej Shevchenko^,||

Max Planck Institute of Molecular Cell Biology and Genetics (MPI CBG)
Genomics, Technische Universität Dresden, c/o MPI CBG, Pfotenhauerstrasse 108, 01307 Dresden, Germany

We employed a combination of tandem affinity purification and mass spectrometry for deciphering protein complexes and the protein interaction network in budding yeast. 53 genes were epitope-tagged, and their interaction partners were isolated by two-step immunoaffinity chromatography from whole cell lysates. 38 baits pulled down a total of 220 interaction partners, which are members of 19 functionally distinct protein complexes. We identified four proteins shared between complexes of different functionality thus charting segments of a protein interaction network. Concordance with the results of genome-wide two-hybrid screening was poor (14% of identified interactors overlapped) suggesting that the two approaches may provide complementary views on physical interactions within the proteome.

^|| To whom correspondence may be addressed. Tel.: 49-351-210-2615; Fax: 49-351-210-2000; E-mail: shevchenko{at}mpi-cbg.de.

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