MilliporeSigma
  • Home
  • Search Results
  • Shiga toxin glycosphingolipid receptors of Vero-B4 kidney epithelial cells and their membrane microdomain lipid environment.

Shiga toxin glycosphingolipid receptors of Vero-B4 kidney epithelial cells and their membrane microdomain lipid environment.

Journal of lipid research (2015-10-16)
Daniel Steil, Catherine-Louise Schepers, Gottfried Pohlentz, Nadine Legros, Jana Runde, Hans-Ulrich Humpf, Helge Karch, Johannes Müthing
ABSTRACT

Shiga toxins (Stxs) are produced by enterohemorrhagic Escherichia coli (EHEC), which cause human infections with an often fatal outcome. Vero cell lines, derived from African green monkey kidney, represent the gold standard for determining the cytotoxic effects of Stxs. Despite their global use, knowledge about the exact structures of the Stx receptor glycosphingolipids (GSLs) and their assembly in lipid rafts is poor. Here we present a comprehensive structural analysis of Stx receptor GSLs and their distribution to detergent-resistant membranes (DRMs), which were prepared from Vero-B4 cells and used as lipid raft equivalents. We identified globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer) as the GSL receptors for Stx1a, Stx2a, and Stx2e subtypes using TLC overlay detection combined with MS. The uncommon Stx receptor, globopentaosylceramide (Gb5Cer, Galβ3GalNAcβ3Galα4Galβ4Glcβ1Cer), which was specifically recognized (in addition to Gb3Cer and Gb4Cer) by Stx2e, was fully structurally characterized. Lipoforms of Stx receptor GSLs were found to mainly harbor ceramide moieties composed of sphingosine (d18:1) and C24:0/C24:1 or C16:0 fatty acid. Moreover, co-occurrence with lipid raft markers, SM and cholesterol, in DRMs suggested GSL association with membrane microdomains. This study provides the basis for further exploring the functional impact of lipid raft-associated Stx receptors for toxin-mediated injury of Vero-B4 cells.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Phenylacetic acid, suitable for plant cell culture
Sigma-Aldrich
5-Bromo-4-chloro-3-indolyl phosphate p-toluidine salt, tablet
Sigma-Aldrich
Sucrose, ≥99.5%
Sigma-Aldrich
5-Bromo-4-chloro-3-indolyl phosphate p-toluidine salt, ≥99% (HPLC)
Sigma-Aldrich
Sucrose, BioReagent, suitable for cell culture, suitable for insect cell culture, ≥99.5% (GC)
Sigma-Aldrich
5-Bromo-4-chloro-3-indolyl phosphate p-toluidine salt, BioReagent, for molecular biology, powder, ≥99%
Supelco
Sucrose, analytical standard
Sigma-Aldrich
Phenylacetic acid, 99%
Sigma-Aldrich
Sucrose, BioUltra, for molecular biology, ≥99.5% (HPLC)
Sigma-Aldrich
5-Bromo-4-chloro-3-indolyl phosphate p-toluidine salt, ≥95.0% (HPLC)
Sigma-Aldrich
Sucrose, puriss., meets analytical specification of Ph. Eur., BP, NF
Sigma-Aldrich
Sucrose, ≥99.5% (GC)
Sigma-Aldrich
Sucrose, BioXtra, ≥99.5% (GC)
Sigma-Aldrich
Sucrose, ACS reagent
Sigma-Aldrich
Sucrose, meets USP testing specifications
Sigma-Aldrich
Sucrose, for molecular biology, ≥99.5% (GC)
Sigma-Aldrich
Phenylacetic acid, ≥99%, FCC, FG
Sigma-Aldrich
Methanol-12C, 99.95 atom % 12C
Supelco
Methanol solution, NMR reference standard, 4% in methanol-d4 (99.8 atom % D), NMR tube size 5 mm × 8 in.
Sigma-Aldrich
Methanol solution, NMR reference standard, 4% in methanol-d4 (99.8 atom % D), NMR tube size 3 mm × 8 in.
SAFC
Cholesterol, Plant-Derived, SyntheChol®, NF, Ph. Eur., JP
Supelco
Methanol solution, contains 0.10 % (v/v) formic acid, UHPLC, suitable for mass spectrometry (MS), ≥99.5%
Sigma-Aldrich
Sucrose, Grade I, suitable for plant cell culture
Sigma-Aldrich
Sucrose, Grade II, suitable for plant cell culture
Sigma-Aldrich
Chloroform, biotech. grade, ≥99.8%, contains 0.5-1.0% ethanol as stabilizer
Sigma-Aldrich
Chloroform, ACS spectrophotometric grade, ≥99.8%, contains 0.5-1.0% ethanol as stabilizer
Sigma-Aldrich
Sodium pyruvate, anhydrous, free-flowing, Redi-Dri, ReagentPlus®, ≥99%
Sigma-Aldrich
Chloroform, ReagentPlus®, ≥99.8%, contains 0.5-1.0% ethanol as stabilizer
Sigma-Aldrich
Chloroform, contains 100-200 ppm amylenes as stabilizer, ≥99.5%
Sigma-Aldrich
Chloroform, contains ethanol as stabilizer, ACS reagent, ≥99.8%