Genetically Encoded Multispectral Labeling of Proteins with Polyfluorophores on a DNA Backbone
- 16 April 2013
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of the American Chemical Society
- Vol. 135 (16), 6184-6191
- https://doi.org/10.1021/ja4004393
Abstract
Genetically encoded methods for protein conjugation are of high importance as biological tools. Here we describe the development of a new class of dyes for genetically encoded tagging that add new capabilities for protein reporting and detection via HaloTag methodology. Oligodeoxyfluorosides (ODFs) are short DNA-like oligomers in which the natural nucleic acid bases are replaced by interacting fluorescent chromophores, yielding a broad range of emission colors using a single excitation wavelength. We describe the development of an alkyl halide dehalogenase-compatible chloroalkane linker phosphoramidite derivative that enables the rapid automated synthesis of many possible dyes for protein conjugation. Experiments to test the enzymatic self-conjugation of nine different DNA-like dyes to proteins with HaloTag domains in vitro were performed, and the data confirmed the rapid and efficient covalent labeling of the proteins. Notably, a number of the ODF dyes were found to increase in brightness or change color upon protein conjugation. Tests in mammalian cellular settings revealed that the dyes are functional in multiple cellular contexts, both on the cell surface and within the cytoplasm, allowing protein localization to be imaged in live cells by epifluorescence and laser confocal microscopy.This publication has 42 references indexed in Scilit:
- Multispectral labeling of antibodies with polyfluorophores on a DNA backbone and application in cellular imagingProceedings of the National Academy of Sciences of the United States of America, 2011
- Differentiating between Fluorescence-Quenching Metal Ions with Polyfluorophore Sensors Built on a DNA BackboneJournal of the American Chemical Society, 2011
- Superresolution Imaging of Targeted Proteins in Fixed and Living Cells Using Photoactivatable Organic FluorophoresJournal of the American Chemical Society, 2010
- Polyfluorophores on a DNA Backbone: Sensors of Small Molecules in the Vapor PhaseAngewandte Chemie, 2010
- A fluorophore ligase for site-specific protein labeling inside living cellsProceedings of the National Academy of Sciences of the United States of America, 2010
- Next-generation quantum dotsNature Biotechnology, 2009
- In Vivo Stable Tumor-Specific Painting in Various Colors Using Dehalogenase-Based Protein-Tag Fluorescent LigandsBioconjugate Chemistry, 2009
- Polyfluorophores on a DNA Backbone: A Multicolor Set of Labels Excited at One WavelengthJournal of the American Chemical Society, 2009
- HaloTag protein-mediated specific labeling of living cells with quantum dotsBiochemical and Biophysical Research Communications, 2008
- Labeling of fusion proteins with synthetic fluorophores in live cellsProceedings of the National Academy of Sciences of the United States of America, 2004