The cell division cycle of the yeast S. cerevisiae is driven by one Cdk (cyclin-dependent kinase), which becomes active when bound to one of nine cyclin subunits. Elucidation of Cdk substrates and other Cdk-associated proteins is essential for a full understanding of the cell cycle. Here, we report the results of a targeted proteomics study using affinity purification coupled to mass spectrometry. Our study identified numerous proteins in association with particular cyclin-Cdk complexes. These included phosphorylation substrates, ubiquitination-degradation proteins, adaptors, and inhibitors. Some associations were previously known, and for others, we confirmed their specificity and biological relevance. Using a hypothesis-driven mass spectrometric approach, we also mapped in vivo phosphorylation at Cdk consensus motif-containing peptides within several cyclin-associated candidate Cdk substrates. Our results demonstrate that this approach can be used to detect a host of transient and dynamic protein associations within a biological module.

• PMID: 15200949
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Journal Article | Research Support, Non-U.S. Gov't | Research Support, U.S. Gov't, P.H.S.

Authors

Archambault V, Chang EJ, Drapkin BJ, Cross FR, Chait BT, Rout MP

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