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 Citations (Scopus)

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
Externally publishedYes

Fingerprint

Fluorescent Dyes
Coloring Agents
Lipids
Pharmaceutical Preparations
Titration
Carrier Proteins
Butyric Acid
Proteins
Wavelength
Calorimetry
Hydrophobicity
Bovine Serum Albumin
Hydrophobic and Hydrophilic Interactions
Solubility
Substitution reactions
Experiments
Binding Sites
Nuclear magnetic resonance
Ligands
Water

Keywords

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

ASJC Scopus subject areas

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

Cite this

Black, S. L., Stanley, W. A., Filipp, F. V., Bhairo, M., Verma, A., Wichmann, O., ... 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

Probing lipid- and drug-binding domains with fluorescent dyes. / Black, Shannon L.; Stanley, Will A.; Filipp, Fabian V.; Bhairo, Michelle; Verma, Ashwani; Wichmann, Oliver; Sattler, Michael; Wilmanns, Matthias; Schultz, Carsten.

In: Bioorganic and Medicinal Chemistry, Vol. 16, No. 3, 01.02.2008, p. 1162-1173.

Research output: Contribution to journalArticle

Black, SL, Stanley, WA, Filipp, FV, 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, vol. 16, no. 3, pp. 1162-1173. https://doi.org/10.1016/j.bmc.2007.10.080
Black SL, Stanley WA, Filipp FV, Bhairo M, Verma A, Wichmann O et al. Probing lipid- and drug-binding domains with fluorescent dyes. Bioorganic and Medicinal Chemistry. 2008 Feb 1;16(3):1162-1173. https://doi.org/10.1016/j.bmc.2007.10.080
Black, Shannon L. ; Stanley, Will A. ; Filipp, Fabian V. ; Bhairo, Michelle ; Verma, Ashwani ; Wichmann, Oliver ; Sattler, Michael ; Wilmanns, Matthias ; Schultz, Carsten. / Probing lipid- and drug-binding domains with fluorescent dyes. In: Bioorganic and Medicinal Chemistry. 2008 ; Vol. 16, No. 3. pp. 1162-1173.
@article{80e67478f11540368acc015f7cec7b27,
title = "Probing lipid- and drug-binding domains with fluorescent dyes",
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.",
keywords = "Binding, Bovine serum albumin, Environmentally sensitive dyes, Isothermal calorimetry, Nile red, SCP2",
author = "Black, {Shannon L.} and Stanley, {Will A.} and Filipp, {Fabian V.} and Michelle Bhairo and Ashwani Verma and Oliver Wichmann and Michael Sattler and Matthias Wilmanns and Carsten Schultz",
year = "2008",
month = "2",
day = "1",
doi = "10.1016/j.bmc.2007.10.080",
language = "English (US)",
volume = "16",
pages = "1162--1173",
journal = "Bioorganic and Medicinal Chemistry",
issn = "0968-0896",
publisher = "Elsevier Limited",
number = "3",

}

TY - JOUR

T1 - Probing lipid- and drug-binding domains with fluorescent dyes

AU - Black, Shannon L.

AU - Stanley, Will A.

AU - Filipp, Fabian V.

AU - Bhairo, Michelle

AU - Verma, Ashwani

AU - Wichmann, Oliver

AU - Sattler, Michael

AU - Wilmanns, Matthias

AU - Schultz, Carsten

PY - 2008/2/1

Y1 - 2008/2/1

N2 - 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.

AB - 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.

KW - Binding

KW - Bovine serum albumin

KW - Environmentally sensitive dyes

KW - Isothermal calorimetry

KW - Nile red

KW - SCP2

UR - http://www.scopus.com/inward/record.url?scp=38849200432&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=38849200432&partnerID=8YFLogxK

U2 - 10.1016/j.bmc.2007.10.080

DO - 10.1016/j.bmc.2007.10.080

M3 - Article

C2 - 18024138

AN - SCOPUS:38849200432

VL - 16

SP - 1162

EP - 1173

JO - Bioorganic and Medicinal Chemistry

JF - Bioorganic and Medicinal Chemistry

SN - 0968-0896

IS - 3

ER -