Red fluorescent protein with reversibly photoswitchable absorbance for photochromic FRET.
We have developed the first red fluorescent protein, named rsTagRFP, which possesses reversibly photoswitchable absorbance spectra. Illumination with blue and yellow light switches rsTagRFP into a red fluorescent state (ON state) or nonfluorescent state (OFF state), respectively. The ON and OFF states exhibit absorbance maxima at 567 and 440 nm, respectively. Due to the photoswitchable absorbance, rsTagRFP can be used as an acceptor for a photochromic Förster resonance energy transfer (pcFRET). The photochromic acceptor facilitates determination of a protein-protein interaction by providing an internal control for FRET. Using pcFRET with EYFP as a donor, we observed an interaction between epidermal growth factor receptor and growth factor receptor-binding protein 2 in live cells by detecting the modulation of both the fluorescence intensity and lifetime of the EYFP donor upon the ON-OFF photoswitching of the rsTagRFP acceptor.
Subach
Fedor V
FV
Department of Anatomy and Structural Biology and Gruss-Lipper Biophotonics Center, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
Zhang
Lijuan
L
Gadella
Theodorus W J
TW
Gurskaya
Nadya G
NG
Lukyanov
Konstantin A
KA
Verkhusha
Vladislav V
VV
D002470:Cell Survival; D066246:ErbB Receptors / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D051380:GRB2 Adaptor Protein / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008027:Light; D008164:Luminescent Proteins / Q000737:chemistry* / Q000235:genetics*; D016296:Mutagenesis; D009154:Mutation; D015202:Protein Engineering / Q000379:methods*; D000097573:Red Fluorescent Protein
D066246:ErbB Receptors; D051380:GRB2 Adaptor Protein; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein