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

Marcus Jäger, Xavier Michalet, Shimon Weiss

Research output: Contribution to journalArticlepeer-review

32 Scopus citations

Abstract

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.

Original languageEnglish
Pages (from-to)2059-2068
Number of pages10
JournalProtein Science
Volume14
Issue number8
DOIs
StatePublished - Aug 2005
Externally publishedYes

Funding

FundersFunder number
National Institute of General Medical SciencesR01GM065382

    Keywords

    • Alternating laser excitation
    • Fluorescence resonance energy transfer (FRET)
    • Fluorescence-aided molecular sorting
    • Protein folding
    • Protein labeling
    • Protein-protein interaction
    • Single molecule spectroscopy

    Fingerprint

    Dive into the research topics of 'Protein-protein interactions as a tool for site-specific labeling of proteins'. Together they form a unique fingerprint.

    Cite this