Probing lipid- and drug-binding domains with fluorescent dyes

Shannon L. Black, Will A. Stanley, Fabian V. Filipp, Michelle Bhairo, Ashwani Verma, Oliver Wichmann, Michael Sattler, Matthias Wilmanns, Carsten Schultz

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

A series of 2- and 3-OH Nile red dyes was prepared in order to generate water-soluble probes that could be used to probe lipid binding to proteins. Various substitutions in positions 2-/3-, 6-, and 7-shifted wavelengths while maintaining the environmental sensitivity of Nile red. In order to increase the solubility of the dyes in aqueous solutions, we attached butyric acid groups to the 2- or 3-OH position. In addition, phenothiazine dyes, which exhibited particularly long excitation properties, were synthesized and tested for the first time. All dyes showed Stoke's shifts of 70-100 nm and changes in excitation and emission of over 100 nm, depending on the hydrophobicity of the environment. Binding studies with bovine serum albumin and the non-specific lipid transfer protein SCP2 revealed emission changes of more than 30 nm upon binding to the protein and a five-fold increase in emission intensity. Titration of the dye-loaded proteins with various lipids or drugs replaced the dye and thereby reversed the shift in wavelength intensity. This allowed us to estimate the lipid binding affinity of the investigated proteins. For SCP2, isothermal calorimetry (ITC) data verified the titration experiments. NMR titration experiments of SCP2 with Nile red 2-O-butyric acid (1a) revealed that the dye is bound within the lipid binding pocket and competes with lipid ligands for this binding site. These results give valuable insight into lipid and drug transport by proteins outside and inside cells.

Original languageEnglish (US)
Pages (from-to)1162-1173
Number of pages12
JournalBioorganic and Medicinal Chemistry
Volume16
Issue number3
DOIs
StatePublished - Feb 1 2008

Keywords

  • Binding
  • Bovine serum albumin
  • Environmentally sensitive dyes
  • Isothermal calorimetry
  • Nile red
  • SCP2

ASJC Scopus subject areas

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

Fingerprint Dive into the research topics of 'Probing lipid- and drug-binding domains with fluorescent dyes'. Together they form a unique fingerprint.

  • Cite this

    Black, S. L., Stanley, W. A., Filipp, F. V., Bhairo, M., Verma, A., Wichmann, O., Sattler, M., Wilmanns, M., & Schultz, C. (2008). Probing lipid- and drug-binding domains with fluorescent dyes. Bioorganic and Medicinal Chemistry, 16(3), 1162-1173. https://doi.org/10.1016/j.bmc.2007.10.080