N-Myristoyl Transferase (NMT)-Catalyzed Labeling of Bacterial Proteins for Imaging in Fixed and Live Cells
| Autor: | Samuel H. Ho, David A. Tirrell |
|---|---|
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
chemistry.chemical_classification Fluorescence-lifetime imaging microscopy Fatty acid 010402 general chemistry medicine.disease_cause 01 natural sciences 0104 chemical sciences 03 medical and health sciences 030104 developmental biology Recognition sequence chemistry Biochemistry Covalent bond Fluorescence microscope medicine Transferase Escherichia coli Myristoylation |
| Zdroj: | Methods in Molecular Biology ISBN: 9781493995455 |
| DOI: | 10.1007/978-1-4939-9546-2_16 |
| Popis: | Methods for selective protein imaging are critical for elucidating how cells orchestrate fundamental biological processes. We recently developed a chemoenzymatic method to modify bacterial proteins in situ for fluorescence imaging using N-myristoyl transferase (NMT). Target proteins outfitted with an N-terminal NMT recognition sequence are covalently modified with an azido fatty acid. Subsequent strain-promoted azide-alkyne cycloaddition allows for conjugation to cell-permeant fluorophores and imaging by fluorescence microscopy. Here we describe sample preparation and labeling protocols for imaging bacterial proteins in fixed and live cells. |
| Databáze: | OpenAIRE |
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