title: Semisynthesis of a protein with cholesterol at the C-terminal, targeted to the cell membrane of live cells.
abstract: Various proteins are modified post-translationally to localize them at the cell membrane. Among them, hedgehog-family proteins are modified by cholesterol at the C-terminal. In this study, green fluorescent protein (GFP) modified with cholesterol (GFP-Chol) at the C-terminal was prepared semisynthetically and investigated. This semi-synthesis was performed using the following native chemical ligation: GFP-Cα-thioester was prepared using the intein-mediated thioester exchange reaction and was ligated to Cys-NH-diethylene glycol-NHCO-cholesterol in the presence of a detergent. After removal of the detergent, the GFP-Chol was applied to mouse live cells. Confocal laser fluorescent microscopy confirmed localization of GFP-Chol at the cell membrane. The findings suggest that modifying proteins with cholesterol at the C-terminal is useful for targeting the proteins to the cell membrane of live cells.
authors.affiliation: Department of Neurochemistry, Tohoku University Graduate School of Medicine, 2-1 Seiryo-Machi, Aoba-Ku, Sendai, Miyagi, 980-8575, Japan.
authors.forename: Kenta
authors.initials: K
authors.lastname: Teruya
authors.forename: Keiko
authors.initials: K
authors.lastname: Nishizawa
authors.forename: Katsumi
authors.initials: K
authors.lastname: Doh-ura
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D002477:Cells / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D002784:Cholesterol / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000138:chemical synthesis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D002784:Cholesterol
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