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Comprehensive Identification of Substrates for F-box Proteins by Differential Proteomics Analysis

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posted on 20.02.2016, 20:41 by Kanae Yumimoto, Masaki Matsumoto, Koji Oyamada, Toshiro Moroishi, Keiichi I. Nakayama
Although elucidation of enzyme–substrate relations is fundamental to the advancement of biology, universal approaches to the identification of substrates for a given enzyme have not been established. It is especially difficult to identify substrates for ubiquitin ligases, given that most such substrates are immediately ubiquitylated and degraded as a result of their association with the enzyme. We here describe the development of a new approach, DiPIUS (differential proteomics-based identification of ubiquitylation substrates), to the discovery of substrates for ubiquitin ligases. We applied DiPIUS to Fbxw7α, Skp2, and Fbxl5, three of the most well-characterized F-box proteins, and identified candidate substrates including previously known targets. DiPIUS is thus a powerful tool for unbiased and comprehensive screening for substrates of ubiquitin ligases.