Synthesis of Lipidated Green Fluorescent Protein and Its Incorporation in Supported Lipid Bilayers

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• 作者：Michael J. Grogan ; Yoshihisa Kaizuka ; Rosemary M. Conrad ; Jay T. Groves ; and Carolyn R. Bertozzi
• 刊名：Journal of the American Chemical Society
• 出版年：2005
• 出版时间：October 19, 2005
• 年：2005
• 卷：127
• 期：41
• 页码：14383 - 14387
• 全文大小：156K
• 年卷期：v.127,no.41(October 19, 2005)
• ISSN：1520-5126

文摘

Herein we report a semisynthetic method of producing membrane-anchored proteins. Ligationof synthetic lipids with designed anchor structures to proteins was performed using native chemical ligation(NCL) of a C-terminal peptide thioester and an N-terminal cysteine lipid. This strategy mimics the naturalglycosylphosphatidylinositol (GPI) linkage found in many natural membrane-associated proteins; however,the synthetic method utilizes simple lipid anchors without glycans. Synthetically lipidated recombinant greenfluorescent protein (GFP) was shown to be stably anchored to the membrane, and its lateral fluidity wasquantitatively characterized by direct fluorescence imaging in supported membranes. Circumventing thesteps of purification from native cell membranes, this methodology facilitates the reconstitution of membrane-associated proteins.