The generality of kinase-catalyzed biotinylation

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• 作者：Chamara Senevirathne ; D. Maheeka Embogama ; Thilani A. Anthony ; Ahmed E. Fouda ; Mary Kay H. Pflum ; ^{pflum@wayne.edu" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Kinase enzymes ; ATP analogs ; Biotinylation ; Kinome tree
• 刊名：Bioorganic & Medicinal Chemistry
• 出版年：2016
• 出版时间：1 January 2016
• 年：2016
• 卷：24
• 期：1
• 页码：12-19
• 全文大小：1583 K

文摘

Kinase-catalyzed protein phosphorylation is involved in a wide variety of cellular events. Development of methods to monitor phosphoproteins in normal and diseased states is critical to fully characterize cell signaling. Towards phosphoprotein analysis tools, our lab reported kinase-catalyzed labeling where γ-phosphate modified ATP analogs are utilized by kinases to label peptides or protein substrates with a functional tag. In particular, the ATP-biotin analog was developed for kinase-catalyzed biotinylation. However, kinase-catalyzed labeling has been tested rigorously with only a few kinases, preventing use of ATP-biotin as a general tool. Here, biotinylation experiments, gel or HPLC-based quantification, and kinetic measurements indicated that twenty-five kinases throughout the kinome tree accepted ATP-biotin as a cosubstrate. With this rigorous characterization of ATP-biotin compatibility, kinase-catalyzed labeling is now immediately useful for studying phosphoproteins and characterizing the role of phosphorylation in various biological events.