Localization of GroEL determined by in vivo incorporation of a fluorescent amino acid

Godefroid Charbon, Jiangyun Wang, Eric Brustad, Peter G. Schultz, Arthur L. Horwich, Christine Jacobs-Wagner, Eli Chapman

Research output: Contribution to journalArticle

13 Scopus citations

Abstract

The molecular chaperone GroEL is required for bacterial growth under all conditions, mediating folding assistance, via its central cavity, to a diverse set of cytosolic proteins; yet the subcellular localization of GroEL remains unresolved. An earlier study, using antibody probing of fixed Escherichia coli cells, indicated colocalization with the cell division protein FtsZ at the cleavage furrow, while a second E. coli study of fixed cells indicated more even distribution throughout the cytoplasm. Here, for the first time, we have examined the spatial distribution of GroEL in living cells using incorporation of a fluorescent unnatural amino acid into the chaperone. Fluorescence microscopy indicated that GroEL is diffusely distributed, both under normal and stress conditions. Importantly, the present procedure uses a small, fluorescent unnatural amino acid to visualize GroEL in vivo, avoiding the steric demands of a fluorescent protein fusion, which compromises proper GroEL assembly. Further, this unnatural amino acid incorporation avoids artifacts that can occur with fixation and antibody staining.

Original languageEnglish (US)
Pages (from-to)6067-6070
Number of pages4
JournalBioorganic and Medicinal Chemistry Letters
Volume21
Issue number20
DOIs
StatePublished - Oct 15 2011
Externally publishedYes

Keywords

  • Chaperone
  • Chaperonin
  • GroEL
  • Protein localization
  • Unnatural amino acid
  • tRNA

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmaceutical Science
  • Drug Discovery
  • Clinical Biochemistry
  • Organic Chemistry

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