Cell-specific Profiling of Nascent Proteomes Using Orthogonal Enzyme-mediated Puromycin Incorporation

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• 作者：Ruth M. Barrett ; Hui-wen Liu ; Haihong Jin ; Richard H. Goodman ; Michael S. Cohen
• 刊名：ACS Chemical Biology
• 出版年：2016
• 出版时间：June 17, 2016
• 年：2016
• 卷：11
• 期：6
• 页码：1532-1536
• 全文大小：280K
• 年卷期：0
• ISSN：1554-8937

文摘

Translation regulation is a fundamental component of gene expression, allowing cells to respond rapidly to a variety of stimuli in the absence of new transcription. The lack of methods for profiling nascent proteomes in distinct cell populations in heterogeneous tissues has precluded an understanding of translational regulation in physiologically relevant contexts. Here, we describe a chemical genetic method that involves orthogonal enzyme-mediated incorporation of a clickable puromycin analogue into nascent polypeptides. Using this method, we show that we can label newly synthesized proteins in a cell-specific manner in cells grown in culture and in heterogeneous tissues. We also show that we can identify the nascent proteome in genetically targeted cell populations using affinity enrichment and tandem mass spectrometry. Our method has the potential to provide unprecedented insights into cell-specific translational regulation in heterogeneous tissues.