Labeling proteins for single-molecule FRET

Chirlmin Joo; Taekjip Ha

doi:10.1101/pdb.prot071035

Labeling proteins for single-molecule FRET

Chirlmin Joo, Taekjip Ha

Department of Physics

Research output: Contribution to journal › Article › peer-review

11 Scopus citations

Abstract

Single-molecule (sm) fluorescence detection is a powerful method for studying biological events without time and population averaging. Förster (fluorescence) resonance energy transfer (FRET) is a spectroscopic technique for measuring distances in the 30-80 Å range in which excitation energy of a donor molecule is transferred to an acceptor via interaction between two induced dipoles. A variant of smFRET is based on total internal reflection (TIR) microscopy. This protocol describes the labeling of protein for smFRET with TIR microscopy. It is based on a labeling procedure for E. coli Rep helicase. A different assay (e.g., different chemical conditions) may be required for other proteins.

Original language	English
Pages (from-to)	1009-1012
Number of pages	4
Journal	Cold Spring Harbor Protocols
Volume	7
Issue number	9
DOIs	https://doi.org/10.1101/pdb.prot071035
State	Published - Sep 2012

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Labeling proteins for single-molecule FRET

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