Optimized fluorescent trimethoprim derivatives for in vivo protein labeling

Nathaniel T. Calloway, Michael Choob, Ana Sanz, Michael Sheetz, Lawrence W. Miller, Virginia W. Cornish

Research output: Contribution to journalArticle

63 Citations (Scopus)

Abstract

The combined technologies of optical microscopy and selective probes allow for real-time analysis of protein function in living cells. Synthetic chemistry offers a means to develop specific, protein-targeted probes that exhibit greater optical and chemical functionality than the widely used fluorescent proteins. Here we describe pharmacokinetically optimized, fluorescent trimethoprim (TMP) analogues that can be used to specifically label recombinant proteins fused to E. coli dihydrofolate reductase (eDHFR) in living, wild-type mammalian cells. These improved fluorescent tags exhibited high specificity and fast labeling kinetics, and they could be detected at a high signal-to-noise ratio by using fluorescence microscopy and fluorescence-activated cell sorting (FACS). We also show that fluorescent TMP-eDHFR complexes are complements to green fluorescent protein (GFP) for two-color protein labeling experiments in cells.

Original languageEnglish (US)
Pages (from-to)767-774
Number of pages8
JournalChemBioChem
Volume8
Issue number7
DOIs
StatePublished - May 7 2007
Externally publishedYes

Fingerprint

Trimethoprim
Labeling
Derivatives
Tetrahydrofolate Dehydrogenase
Cells
Escherichia coli
Proteins
Fluorescence microscopy
Signal-To-Noise Ratio
Green Fluorescent Proteins
Fluorescence Microscopy
Sorting
Recombinant Proteins
Optical microscopy
Labels
Microscopy
Signal to noise ratio
Flow Cytometry
Color
Fluorescence

Keywords

  • Bioorganic chemistry
  • Fluorescent probes
  • Green fluorescent protein
  • Protein modifications
  • Trimethoprim

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Organic Chemistry

Cite this

Calloway, N. T., Choob, M., Sanz, A., Sheetz, M., Miller, L. W., & Cornish, V. W. (2007). Optimized fluorescent trimethoprim derivatives for in vivo protein labeling. ChemBioChem, 8(7), 767-774. https://doi.org/10.1002/cbic.200600414

Optimized fluorescent trimethoprim derivatives for in vivo protein labeling. / Calloway, Nathaniel T.; Choob, Michael; Sanz, Ana; Sheetz, Michael; Miller, Lawrence W.; Cornish, Virginia W.

In: ChemBioChem, Vol. 8, No. 7, 07.05.2007, p. 767-774.

Research output: Contribution to journalArticle

Calloway, NT, Choob, M, Sanz, A, Sheetz, M, Miller, LW & Cornish, VW 2007, 'Optimized fluorescent trimethoprim derivatives for in vivo protein labeling', ChemBioChem, vol. 8, no. 7, pp. 767-774. https://doi.org/10.1002/cbic.200600414
Calloway, Nathaniel T. ; Choob, Michael ; Sanz, Ana ; Sheetz, Michael ; Miller, Lawrence W. ; Cornish, Virginia W. / Optimized fluorescent trimethoprim derivatives for in vivo protein labeling. In: ChemBioChem. 2007 ; Vol. 8, No. 7. pp. 767-774.
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