Journal ArticleDOI
Specific Covalent Labeling of Recombinant Protein Molecules Inside Live Cells
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TLDR
This system provides a recipe for slightly modifying a target protein so that it can be singled out from the many other proteins inside live cells and fluorescently stained by small nonfluorescent dye molecules added from outside the cells.Abstract:
Recombinant proteins containing four cysteines at the i , i + 1, i + 4, and i + 5 positions of an α helix were fluorescently labeled in living cells by extracellular administration of 4′,5′-bis(1,3,2-dithioarsolan-2-yl)fluorescein. This designed small ligand is membrane-permeant and nonfluorescent until it binds with high affinity and specificity to the tetracysteine domain. Such in situ labeling adds much less mass than does green fluorescent protein and offers greater versatility in attachment sites as well as potential spectroscopic and chemical properties. This system provides a recipe for slightly modifying a target protein so that it can be singled out from the many other proteins inside live cells and fluorescently stained by small nonfluorescent dye molecules added from outside the cells.read more
Citations
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Fluorescence as a method to reveal structures and membrane-interactions of amyloidogenic proteins.
TL;DR: A review of investigations of the interactions of amyloidogenic polypeptides and their aggregated states with membranes highlights the significant role played by fluorescence techniques in unraveling the nature ofAmyloid fibrils and their interactions and effects on membranes.
Journal ArticleDOI
Quantitative four-dimensional tracking of cytoplasmic and nuclear HIV-1 complexes
Nathalie J. Arhel,Auguste Genovesio,Kyeong-Ae Kim,Sarah Miko,Emmanuelle Perret,Jean-Christophe Olivo-Marin,Spencer L. Shorte,Pierre Charneau +7 more
TL;DR: This work labeled human immunodeficiency virus-1 (HIV-1) integrase with a small tetracysteine tag, which preserved the virus' infectivity while allowing it to be labeled with the bis-arsenical fluorescein derivative FlAsH, which allowed this work to image both intracytoplasmic and intranuclear HIV-1 complexes in three dimensions over time (4D) in human cells and enabled the study of virus-host
Journal ArticleDOI
Reversible site-selective labeling of membrane proteins in live cells.
TL;DR: The ionotropic 5HT3 serotonin receptor is investigated by fluorescence measurements to characterize in vivo the probe-receptor interactions, yielding information on structure and plasma membrane distribution of the receptor.
Journal ArticleDOI
In vivo protein labeling with trimethoprim conjugates: a flexible chemical tag
TL;DR: It is demonstrated that trimethoprim derivatives can be used to selectively tag Escherichia coli dihydrofolate reductase (eDHFR) fusion proteins in wild-type mammalian cells with minimal background and fast kinetics.
Journal ArticleDOI
Organelle-Targetable Fluorescent Probes for Imaging Hydrogen Peroxide in Living Cells via SNAP-Tag Protein Labeling
TL;DR: This work has exploited the versatility of the SNAP-tag technology for site-specific protein labeling with small molecules on the surface or interior of living cells with the use of boronate-capped dyes to selectively visualize H(2)O(2).
References
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Journal ArticleDOI
The green fluorescent protein
TL;DR: In just three years, the green fluorescent protein from the jellyfish Aequorea victoria has vaulted from obscurity to become one of the most widely studied and exploited proteins in biochemistry and cell biology.
Journal ArticleDOI
Rapid colorimetric assay for cell growth and survival. Modifications to the tetrazolium dye procedure giving improved sensitivity and reliability.
François Denizot,Rita Lang +1 more
TL;DR: The reliability and sensitivity of the test have been increased to the point where it can in many cases replace the [3H]thymidine uptake assay to measure cell proliferation or survival in growth factor or cytotoxicity assays.
Journal ArticleDOI
Fluorescent indicators for Ca2+based on green fluorescent proteins and calmodulin
Atsushi Miyawaki,Juan Llopis,Roger Heim,J. Michael McCaffery,Joseph A. Adams,Mitsuhiko Ikura,Mitsuhiko Ikura,Roger Y. Tsien +7 more
TL;DR: New fluorescent indicators for Ca2+ that are genetically encoded without cofactors and are targetable to specific intracellular locations are constructed and dubbed ‘cameleons’.
Journal ArticleDOI
Crystal structure of the Aequorea victoria green fluorescent protein.
TL;DR: The green fluorescent protein (GFP) from the Pacific Northwest jellyfish Aequorea victoria has generated intense interest as a marker for gene expression and localization of gene products.
Journal ArticleDOI
Engineering green fluorescent protein for improved brightness, longer wavelengths and fluorescence resonance energy transfer
Roger Heim,Roger Y. Tsien +1 more
TL;DR: The results demonstrate that the production of more and better GFP variants is possible and worthwhile, and facilitates multicolor imaging of differential gene expression, protein localization or cell fate.
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