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A FRET-facilitated photoswitching using an orange fluorescent protein with the fast photoconversion kinetics.
J Am Chem Soc 2012; 134(36):14789-99JA

Abstract

Fluorescent proteins photoswitchable with noncytotoxic light irradiation and spectrally distinct from multiple available photoconvertible green-to-red probes are in high demand. We have developed a monomeric fluorescent protein, called PSmOrange2, which is photoswitchable with blue light from an orange (ex./em. at 546 nm/561 nm) to a far-red (ex./em. at 619 nm/651 nm) form. Compared to another orange-to-far-red photoconvertable variant, PSmOrange2 has blue-shifted photoswitching action spectrum, 9-fold higher photoconversion contrast, and up to 10-fold faster photoswitching kinetics. This results in the 4-fold more PSmOrange2 molecules being photoconverted in mammalian cells. Compared to common orange fluorescent proteins, such as mOrange, the orange form of PSmOrange has substantially higher photostability allowing its use in multicolor imaging applications to track dynamics of multiple populations of intracellular objects. The PSmOrange2 photochemical properties allow its efficient photoswitching with common two-photon lasers and, moreover, via Förster resonance energy transfer (FRET) from green fluorescent donors. We have termed the latter effect a FRET-facilitated photoswitching and demonstrated it using several sets of interacting proteins. The enhanced photoswitching properties of PSmOrange2 make it a superior photoconvertable protein tag for flow cytometry, conventional microscopy, and two-photon imaging of live cells.

Authors+Show Affiliations

Department of Anatomy and Structural Biology and Gruss-Lipper Biophotonics Center, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural

Language

eng

PubMed ID

22900938

Citation

Subach, Oksana M., et al. "A FRET-facilitated Photoswitching Using an Orange Fluorescent Protein With the Fast Photoconversion Kinetics." Journal of the American Chemical Society, vol. 134, no. 36, 2012, pp. 14789-99.
Subach OM, Entenberg D, Condeelis JS, et al. A FRET-facilitated photoswitching using an orange fluorescent protein with the fast photoconversion kinetics. J Am Chem Soc. 2012;134(36):14789-99.
Subach, O. M., Entenberg, D., Condeelis, J. S., & Verkhusha, V. V. (2012). A FRET-facilitated photoswitching using an orange fluorescent protein with the fast photoconversion kinetics. Journal of the American Chemical Society, 134(36), pp. 14789-99. doi:10.1021/ja3034137.
Subach OM, et al. A FRET-facilitated Photoswitching Using an Orange Fluorescent Protein With the Fast Photoconversion Kinetics. J Am Chem Soc. 2012 Sep 12;134(36):14789-99. PubMed PMID: 22900938.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A FRET-facilitated photoswitching using an orange fluorescent protein with the fast photoconversion kinetics. AU - Subach,Oksana M, AU - Entenberg,David, AU - Condeelis,John S, AU - Verkhusha,Vladislav V, Y1 - 2012/08/27/ PY - 2012/8/21/entrez PY - 2012/8/21/pubmed PY - 2013/2/5/medline SP - 14789 EP - 99 JF - Journal of the American Chemical Society JO - J. Am. Chem. Soc. VL - 134 IS - 36 N2 - Fluorescent proteins photoswitchable with noncytotoxic light irradiation and spectrally distinct from multiple available photoconvertible green-to-red probes are in high demand. We have developed a monomeric fluorescent protein, called PSmOrange2, which is photoswitchable with blue light from an orange (ex./em. at 546 nm/561 nm) to a far-red (ex./em. at 619 nm/651 nm) form. Compared to another orange-to-far-red photoconvertable variant, PSmOrange2 has blue-shifted photoswitching action spectrum, 9-fold higher photoconversion contrast, and up to 10-fold faster photoswitching kinetics. This results in the 4-fold more PSmOrange2 molecules being photoconverted in mammalian cells. Compared to common orange fluorescent proteins, such as mOrange, the orange form of PSmOrange has substantially higher photostability allowing its use in multicolor imaging applications to track dynamics of multiple populations of intracellular objects. The PSmOrange2 photochemical properties allow its efficient photoswitching with common two-photon lasers and, moreover, via Förster resonance energy transfer (FRET) from green fluorescent donors. We have termed the latter effect a FRET-facilitated photoswitching and demonstrated it using several sets of interacting proteins. The enhanced photoswitching properties of PSmOrange2 make it a superior photoconvertable protein tag for flow cytometry, conventional microscopy, and two-photon imaging of live cells. SN - 1520-5126 UR - https://www.unboundmedicine.com/medline/citation/22900938/A_FRET_facilitated_photoswitching_using_an_orange_fluorescent_protein_with_the_fast_photoconversion_kinetics_ L2 - https://dx.doi.org/10.1021/ja3034137 DB - PRIME DP - Unbound Medicine ER -