期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 105, 期 47, 页码 18343-18348出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0805949105
关键词
fluorescent proteins; microspectrophotometry; photoactivation; photochromism; protein crystallography
资金
- Agence Nationale de la Recherche [ANR-07-BLAN-0107-01]
- Deutsche Forschungsgemeinschaft [N1 291/9, SFB 497]
- Fonds der Chemischen Industrie
- Landesstiftung Baden-Wurttemberg
- Agence Nationale de la Recherche (ANR) [ANR-07-BLAN-0107] Funding Source: Agence Nationale de la Recherche (ANR)
Photoactivatable fluorescent proteins (FPs) are powerful fluorescent highlighters in live cell imaging and offer perspectives for optical nanoscopy and the development of biophotonic devices. Two types of photoactivation are currently being distinguished, reversible photoswitching between fluorescent and nonfluorescent forms and irreversible photoconversion. Here, we have combined crystallography and (in crystallo) spectroscopy to characterize the Phe-173-Ser mutant of the tetrameric variant of EosFP, named IrisFP, which incorporates both types of phototransformations. In its green fluorescent state, IrisFP displays reversible photoswitching, which involves cis-trans isomerization of the chromophore. Like its parent protein EosFP, IrisFP also photoconverts irreversibly to a red-emitting state under violet light because of an extension of the conjugated pi-electron system of the chromophore, accompanied by a cleavage of the polypeptide backbone. The red form of IrisFP exhibits a second reversible photoswitching process, which may also involve cis-trans isomerization of the chromophore. Therefore, IrisFP displays altogether 3 distinct photoactivation processes. The possibility to engineer and precisely control multiple phototransformations in photoactivatable FPs offers exciting perspectives for the extension of the fluorescent protein toolkit.
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