Abstract
We report a photoswitchable monomeric Orange (PSmOrange) protein that is initially orange (excitation, 548 nm; emission, 565 nm) but becomes far-red (excitation, 636 nm; emission, 662 nm) after irradiation with blue-green light. Compared to its parental orange proteins, PSmOrange has greater brightness, faster maturation, higher photoconversion contrast and better photostability. The red-shifted spectra of both forms of PSmOrange enable its simultaneous use with cyan-to-green photoswitchable proteins to study four intracellular populations. Photoconverted PSmOrange has, to our knowledge, the most far-red excitation peak of all GFP-like fluorescent proteins, provides diffraction-limited and super-resolution imaging in the far-red light range, is optimally excited with common red lasers, and can be photoconverted subcutaneously in a mouse. PSmOrange photoswitching occurs via a two-step photo-oxidation process, which causes cleavage of the polypeptide backbone. The far-red fluorescence of photoconverted PSmOrange results from a new chromophore containing N-acylimine with a co-planar carbon-oxygen double bond.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 771-780 |
| Number of pages | 10 |
| Journal | Nature Methods |
| Volume | 8 |
| Issue number | 9 |
| DOIs | |
| State | Published - Sep 2011 |
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ASJC Scopus subject areas
- Biotechnology
- Molecular Biology
- Biochemistry
- Cell Biology
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A photoswitchable orange-to-far-red fluorescent protein, PSmOrange. / Subach, Oksana M.; Patterson, George H.; Ting, Li Min; Wang, Yarong; Condeelis, John S.; Verkhusha, Vladislav.
In: Nature Methods, Vol. 8, No. 9, 09.2011, p. 771-780.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - A photoswitchable orange-to-far-red fluorescent protein, PSmOrange
AU - Subach, Oksana M.
AU - Patterson, George H.
AU - Ting, Li Min
AU - Wang, Yarong
AU - Condeelis, John S.
AU - Verkhusha, Vladislav
PY - 2011/9
Y1 - 2011/9
N2 - We report a photoswitchable monomeric Orange (PSmOrange) protein that is initially orange (excitation, 548 nm; emission, 565 nm) but becomes far-red (excitation, 636 nm; emission, 662 nm) after irradiation with blue-green light. Compared to its parental orange proteins, PSmOrange has greater brightness, faster maturation, higher photoconversion contrast and better photostability. The red-shifted spectra of both forms of PSmOrange enable its simultaneous use with cyan-to-green photoswitchable proteins to study four intracellular populations. Photoconverted PSmOrange has, to our knowledge, the most far-red excitation peak of all GFP-like fluorescent proteins, provides diffraction-limited and super-resolution imaging in the far-red light range, is optimally excited with common red lasers, and can be photoconverted subcutaneously in a mouse. PSmOrange photoswitching occurs via a two-step photo-oxidation process, which causes cleavage of the polypeptide backbone. The far-red fluorescence of photoconverted PSmOrange results from a new chromophore containing N-acylimine with a co-planar carbon-oxygen double bond.
AB - We report a photoswitchable monomeric Orange (PSmOrange) protein that is initially orange (excitation, 548 nm; emission, 565 nm) but becomes far-red (excitation, 636 nm; emission, 662 nm) after irradiation with blue-green light. Compared to its parental orange proteins, PSmOrange has greater brightness, faster maturation, higher photoconversion contrast and better photostability. The red-shifted spectra of both forms of PSmOrange enable its simultaneous use with cyan-to-green photoswitchable proteins to study four intracellular populations. Photoconverted PSmOrange has, to our knowledge, the most far-red excitation peak of all GFP-like fluorescent proteins, provides diffraction-limited and super-resolution imaging in the far-red light range, is optimally excited with common red lasers, and can be photoconverted subcutaneously in a mouse. PSmOrange photoswitching occurs via a two-step photo-oxidation process, which causes cleavage of the polypeptide backbone. The far-red fluorescence of photoconverted PSmOrange results from a new chromophore containing N-acylimine with a co-planar carbon-oxygen double bond.
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UR - http://www.scopus.com/inward/citedby.url?scp=80052281149&partnerID=8YFLogxK
U2 - 10.1038/nmeth.1664
DO - 10.1038/nmeth.1664
M3 - Article
C2 - 21804536
AN - SCOPUS:80052281149
VL - 8
SP - 771
EP - 780
JO - Nature Methods
JF - Nature Methods
SN - 1548-7091
IS - 9
ER -