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FISHING FOR COMPLEXES
Protein baiting, mass spectrometry allow large-scale analysis of yeast proteome
Sequencing a genome is just the first step in understanding the biology of an organism. Proteins really do the work. With that in mind, two groups independently report similar methods for discovering protein complexes within yeast.
Both groups use "bait" proteins to fish out protein interactions. Yeast proteins are turned into bait by attaching a tag that allows them to be captured in an immunoaffinity purification. The captured complexes are then separated by gel electrophoresis, and excised gel spots are analyzed by mass spectrometry (MS).
A team of scientists from MDS Proteomics in Toronto and Odense, Denmark; Mount Sinai Hospital in Toronto; and the University of Toronto used the method to detect 3,617 protein interactions, starting with 725 bait proteins [Nature, 415, 180 (2002)].
The other team, led by Giulio Superti-Furga, vice president of biology at Cellzome AG and team leader at the European Molecular Biology Laboratory, both in Heidelberg, Germany, identified 232 distinct multiprotein complexes [Nature, 415, 141 (2002)]. They proposed cellular roles for 344 proteins, including 231 for which no function was previously known. In addition to assigning proteins to complexes, the Cellzome scientists investigated relationships between those complexes.
Ruedi Aebersold, a faculty member at the Institute for Systems Biology in Seattle who also uses MS to study proteins, points out that the groups' methods are not new. "It is the amount rather than the type of data that is unusual," Aebersold says, adding that the work is significant because "they are for the first time doing large-scale analysis of protein interactions using a method that is not a genetic method."
Such genetic methods include the yeast two-hybrid method, which identifies pairs of interacting proteins. The techniques in the Nature papers answer different questions than the yeast two-hybrid method, says Daniel Figeys, vice president of analytical sciences at MDS Proteomics.
"The main question that yeast two-hybrid attempts to answer is: 'Is protein A binding to protein B?' Our approach answers the questions: 'What is the identity of all the proteins binding to protein A?' and 'What are the different complexes that protein A is involved in?' and 'What pathways is protein A involved in?' with no prior knowledge of the interacting partners of protein A," Figeys says.
"The network of complexes that we created [provides] an unprecedented view of the proteome at a level of organization beyond individual protein interactions, creating the basis for a reconstruction of the cellular complexity from its components," Superti-Furga says.
The Cellzome study includes examples of complexes from human cells that are remarkably similar to their yeast counterparts. "Just imagine what this could mean for drug discovery," Superti-Furga continues. "With this technology we can elucidate all the molecular machines to which the drug target belongs, offering the prospect of more precise and efficient intervention strategies."
REPRINTED WITH PERMISSION FROM NATURE 2002
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