Conversion of the monomeric red fluorescent protein into a photoactivatable probe

Vladislav Verkhusha, Alexander Sorkin

Research output: Contribution to journalArticle

93 Citations (Scopus)

Abstract

Photoactivatable fluorescent proteins bring new dimension to the analysis of protein dynamics in the cell. Protein tagged with a photoactivatable label can be visualized and tracked in a spatially and temporally defined manner. Here, we describe a basic rational design strategy to develop monomeric photoactivatable proteins using site-specific mutagenesis of common monomeric red-shifted fluorescent proteins. This strategy was applied to mRFP1, which was converted into probes that are photoactivated by either green or violet light. The latter photoactivatable variants, named PA-mRFP1s, exhibited a 70-fold increase of fluorescence intensity resulting from the photoconversion of a violet-light-absorbing precursor. Detailed characterization of PA-mRFP1s was performed with the purified proteins and the proteins expressed in mammalian cells where the photoactivatable properties were preserved. PA-mRFP1s were used as protein tags to study the intracellular dynamics of GTPase Rab5.

Original languageEnglish (US)
Pages (from-to)279-285
Number of pages7
JournalChemistry and Biology
Volume12
Issue number3
DOIs
StatePublished - Mar 2005
Externally publishedYes

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Proteins
Viola
Light
GTP Phosphohydrolases
Mutagenesis
Site-Directed Mutagenesis
red fluorescent protein
Fluorescence
Labels
Cells
Green Or

ASJC Scopus subject areas

  • Organic Chemistry

Cite this

Conversion of the monomeric red fluorescent protein into a photoactivatable probe. / Verkhusha, Vladislav; Sorkin, Alexander.

In: Chemistry and Biology, Vol. 12, No. 3, 03.2005, p. 279-285.

Research output: Contribution to journalArticle

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