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

Herein we report a semisynthetic method of producing membrane-anchored proteins. Ligation of 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 na...

Full description

Saved in:
Bibliographic Details
Published in:Journal of the American Chemical Society 2005-10, Vol.127 (41), p.14383-14387
Main Authors: Grogan, Michael J, Kaizuka, Yoshihisa, Conrad, Rosemary M, Groves, Jay T, Bertozzi, Carolyn R
Format: Article
Language:English
Subjects:
Artificial membranes and reconstituted systems
Biological and medical sciences
Fundamental and applied biological sciences. Psychology
Green Fluorescent Proteins - chemical synthesis
Green Fluorescent Proteins - chemistry
Lipid Bilayers - chemical synthesis
Lipid Bilayers - chemistry
Lipids - chemistry
Membrane physicochemistry
Molecular biophysics
Molecular Structure
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Herein we report a semisynthetic method of producing membrane-anchored proteins. Ligation of 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 natural glycosylphosphatidylinositol (GPI) linkage found in many natural membrane-associated proteins; however, the synthetic method utilizes simple lipid anchors without glycans. Synthetically lipidated recombinant green fluorescent protein (GFP) was shown to be stably anchored to the membrane, and its lateral fluidity was quantitatively characterized by direct fluorescence imaging in supported membranes. Circumventing the steps of purification from native cell membranes, this methodology facilitates the reconstitution of membrane-associated proteins.
ISSN:0002-7863
1520-5126
DOI:10.1021/ja052407f