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Protein–protein interactions as a tool for site-specific labeling of proteins

By Marcus Jäger, Xavier Michalet and Shimon Weiss


Probing structures and dynamics within biomolecules using ensemble and single-molecule fluorescence resonance energy transfer requires the conjugation of fluorophores to proteins in a site-specific and thermodynamically nonperturbative fashion. Using single-molecule fluorescence-aided molecular sorting and the chymotrypsin inhibitor 2–subtilisin BPN′ complex as an example, we demonstrate that protein–protein interactions can be exploited to afford site-specific labeling of a recombinant double-cysteine variant of CI2 without the need for extensive and time-consuming chromatography. The use of protein–protein interactions for site-specific labeling of proteins is compatible with and complementary to existing chemistries for selective labeling of N-terminal cysteines, and could be extended to label multiple positions within a given polypeptide chain

Topics: Article
Publisher: Cold Spring Harbor Laboratory Press
OAI identifier: oai:pubmedcentral.nih.gov:2279317
Provided by: PubMed Central
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