[NMR paper] Rational Engineering of Photoconvertible Fluorescent Proteins for Dual-Color Fluorescence Nanoscopy Enabled by a Triplet-State Mechanism of Primed Conversion
Rational Engineering of Photoconvertible Fluorescent Proteins for Dual-Color Fluorescence Nanoscopy Enabled by a Triplet-State Mechanism of Primed Conversion
Green-to-red photoconvertible fluorescent proteins (pcFPs) are powerful tools for super-resolution localization microscopy and protein tagging. Recently, they have been found to undergo efficient photoconversion not only by the traditional 400-nm illumination but also by an alternative method termed primed conversion, employing dual wavelength illumination with blue and far-red/near-infrared light. Primed conversion has been reported only for Dendra2 and its mechanism has remained elusive. Here, we uncover the molecular mechanism of primed conversion by reporting the intermediate “primed” state to be a triplet dark state formed by intersystem crossing. We show that formation of this state can be influenced by the introduction of serine or threonine at sequence position 69 (Eos notation) and use this knowledge to create “pr”- (for primed convertible) variants of most known green-to-red pcFPs.Uncovering the mechanism of primed conversion allows for rational engineering of primed conversion capable (pr-) fluorescent proteins. An engineered pr-Eos and a wild-type Eos protein are combined for dual-color fluorescence nanoscopy using photoactivated localization microscopy.
[NMR paper] A triplet state mechanism of primed conversion enables rational engineering of photoconvertible fluorescent proteins for dual color fluorescence nanoscopy
A triplet state mechanism of primed conversion enables rational engineering of photoconvertible fluorescent proteins for dual color fluorescence nanoscopy
Photoconvertible fluorescent proteins (pcFPs) are powerful tools for super-resolution localization microscopy and protein tagging. Recently, they have been found to undergo efficient photoconversion by an alternative method termed primed conversion, employing dual wavelength illumination with blue and far-red/near-infrared light. Primed conversion has been reported only for Dendra2 and its mechanism has remained elusive. Here, we...
nmrlearner
Journal club
0
06-30-2017 05:34 AM
[NMR paper] A general mechanism of photoconversion of green-to-red fluorescent proteins based on blue and infrared light reduces phototoxicity in live-cell single-molecule imaging
A general mechanism of photoconversion of green-to-red fluorescent proteins based on blue and infrared light reduces phototoxicity in live-cell single-molecule imaging
Photoconversion of fluorescent proteins by blue and complementary near-infrared light, termed primed conversion (PC), is a mechanism recently discovered for Dendra2. We demonstrate that controlling the conformation of arginine at residue 66 by threonine at residue 69 of fluorescent proteins from Anthozoan families (Dendra2, mMaple, Eos, mKikGR, pcDronpa protein families) represents a general route to facilitate PC....
[NMR paper] Bioconjugation of proteins with a paramagnetic NMR and fluorescent tag.
Bioconjugation of proteins with a paramagnetic NMR and fluorescent tag.
Related Articles Bioconjugation of proteins with a paramagnetic NMR and fluorescent tag.
Chemistry. 2013 Dec 9;19(50):17141-9
Authors: Huang F, Pei YY, Zuo HH, Chen JL, Yang Y, Su XC
Abstract
Site-specific labeling of proteins with lanthanide ions offers great opportunities for investigating the structure, function, and dynamics of proteins by virtue of the unique properties of lanthanides. Lanthanide-tagged proteins can be studied by NMR, X-ray, fluorescence, and EPR...
nmrlearner
Journal club
0
12-07-2013 01:00 PM
Scientists move toward rational design of artificial proteins - R & D Magazine
Scientists move toward rational design of artificial proteins - R & D Magazine
<img alt="" height="1" width="1" />
Scientists move toward rational design of artificial proteins
R & D Magazine
Less vulnerable to chemical or metabolic breakdown than proteins, peptoids are promising for diagnostics, pharmaceuticals, and as a platform to build bioinspired nanomaterials, as scientists can build and manipulate peptoids with great precision. But ...
Read here
nmrlearner
Online News
0
08-23-2012 03:46 AM
[NMR900 blog] Solid-State NMR in Materials for Energy Storage and Conversion
Solid-State NMR in Materials for Energy Storage and Conversion
A special issue of Solid State Nuclear Magnetic Resonance guest-edited by Clare Grey (Cambridge) and Gillian Goward (McMaster), Volume 42, Pages 1-98 (April 2012) http://www.sciencedirect.com/science/journal/09262040/42
Editorial
Clare P. Grey, Gillian R. Goward, Editorial "Solid-State NMR in Materials for Energy Storage and Conversion," Solid State Nuclear Magnetic Resonance
42 (2012) 1. http://dx.doi.org/10.1016/j.ssnmr.2012.03.001
Canadian contributions
nmrlearner
News from NMR blogs
0
03-21-2012 11:40 PM
Systematic Study of Protein Detection Mechanism of Self-Assembling 19F NMR/MRI Nanoprobes toward Rational Design and Improved Sensitivity
Systematic Study of Protein Detection Mechanism of Self-Assembling 19F NMR/MRI Nanoprobes toward Rational Design and Improved Sensitivity
Yousuke Takaoka, Keishi Kiminami, Keigo Mizusawa, Kazuya Matsuo, Michiko Narazaki, Tetsuya Matsuda and Itaru Hamachi
http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jacsat/0/jacsat.ahead-of-print/ja203996c/aop/images/medium/ja-2011-03996c_0004.gif
Journal of the American Chemical Society
DOI: 10.1021/ja203996c
http://feeds.feedburner.com/~ff/acs/jacsat?d=yIl2AUoC8zA
http://feeds.feedburner.com/~r/acs/jacsat/~4/fqTSjFalrGg
Rational Engineering of Photoconvertible Fluorescent Proteins for Dual-Color Fluorescence Nanoscopy Enabled by a Triplet-State Mechanism of Primed Conversion
Linear Mode
