Fluorescent leukotriene B-4: potential applications

Research output: Contribution to journalArticle

Abstract

Leukotriene B-4 (LTB4) is a potent lipid mediator of inflammation that acts primarily via a seven-transmembrane-spanning, G-protein-coupled receptor denoted BLT1. Here, we describe the synthesis and characterization of fluorescent analogs of LTB4 that are easy to produce, inexpensive, and without the disadvantages of a radioligand. Fluorescent LTB4 is useful for labeling LTB4 receptors for which no antibodies are available and for performing one-step fluorescence polarization assays conducive to high-throughput screening. We found that orange and green fluorescent LTB4 were full agonists that activated the LTB4 receptor BLT1 with EC50 values of 68 and 40 nM, respectively (4.5 nM for unmodified LTB4). Flow cytometric measurements and confocal imaging showed that fluorescent LTB4 colocalized with BLT1. Fluorescence polarization measurements showed that orange fluorescent LTB4 bound to BLT1 with a K-d of 66 nM and that this binding could be displaced by unlabeled LTB4 and other BLT1-specific ligands. Fluorescent LTB4 analogs were also able to displace tritiated LTB4. Orange fluorescent LTB4 binding to enhanced green fluorescent protein-tagged BLT1 could be observed using fluorescence resonance energy transfer. In addition to being a useful alternative to radiolabeled LTB4, the unique properties of fluorescently labeled LTB4 allow a variety of detection technologies to be used.

Details

Authors
  • A Sabirsh
  • A Wetterholm
  • Jesper Bristulf
  • Hakon Leffler
  • JZ Haeggstrom
  • Christer Owman
Organisations
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Neurosciences
  • Microbiology in the medical area
  • Immunology in the medical area

Keywords

  • G-protein coupled receptor, pharmacology, method
Original languageEnglish
Pages (from-to)1339-1346
JournalJournal of Lipid Research
Volume46
Issue number6
Publication statusPublished - 2005
Publication categoryResearch
Peer-reviewedYes

Bibliographic note

The information about affiliations in this record was updated in December 2015. The record was previously connected to the following departments: Molecular Neurobiology (0131000140), Division of Microbiology, Immunology and Glycobiology - MIG (013025200), LU Innovation System (018009040), Drug Target Discovery (013212045)