Lipid components of cells and tissues feature a large diversity of structures that present a challenging problem for molecular analysis. Glycolipids from mammalian cells contain glycosphingolipids (GSLs) as their major glycolipid component, and these structures vary in the identity of the glycan headgroup as well as the structure of the fatty acid and sphingosine (Sph) tails. Analysis of intact GSLs is challenging due to the low abundance of these species. Here, we develop a new strategy for the analysis of lyso-GSL (l-GSL), GSL that retain linkage of the glycan headgroup with the Sph base. The analysis begins with digestion of a GSL sample with sphingolipid ceramide N-deacylase (SCDase), followed by labelling with an amine-reactive fluorophore. The sample was then analyzed by HPLC-FLD-MS and quantitated by addition of an external standard. This method was compared to analysis of GSL glycans after cleavage by an Endoglycoceramidase (EGCase) enzyme and labeling with a fluorophore (2-anthranilic acid, 2AA). The two methods are complementary, with EGCase providing improved signal (due to fewer species) and SCDase providing analysis of lyso-GSL. Importantly the SCDase method provides Sph composition of GSL species. We demonstrate the method on cultured human cells (Jurkat T cells) and tissue homogenate (porcine brain).
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1016/j.ab.2021.114361 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Polymyxin B1 in the Escherichia coli inner membrane: A complex story of protein and lipopolysaccharide-mediated insertion.
J Biol Chem
October 2024
Department of Biochemistry, University of Oxford, Porto Alegre, UK. Electronic address:
The rise in multi-drug resistant Gram-negative bacterial infections has led to an increased need for "last-resort" antibiotics such as polymyxins. However, the emergence of polymyxin-resistant strains threatens to bring about a post-antibiotic era. Thus, there is a need to develop new polymyxin-based antibiotics, but a lack of knowledge of the mechanism of action of polymyxins hinders such efforts.
View Article and Find Full Text PDFFAIMS Shotgun Lipidomics for Enhanced Class- and Charge-State Separation Complemented by Automated Ganglioside Annotation.
Anal Chem
July 2024
Department of Analytical Chemistry, Faculty of Chemistry, University of Vienna, Vienna 1090, Austria.
The analysis of gangliosides is extremely challenging, given their structural complexity, lack of reference standards, databases, and software solutions. Here, we introduce a fast 6 min high field asymmetric ion mobility spectrometry (FAIMS) shotgun lipidomics workflow, along with a dedicated software solution for ganglioside detection. By ramping FAIMS compensation voltages, ideal ranges for different ganglioside classes were obtained.
View Article and Find Full Text PDFInteraction of chondroitin sulfate with zwitterionic lipid membranes.
Chem Phys Lipids
September 2024
Jagiellonian University, Faculty of Chemistry, Gronostajowa 2, Kraków 30-387, Poland. Electronic address:
Chondroitin sulfates (CSs) are important components of the extracellular matrix and side chains of membrane proteoglycans. These polysaccharides are, therefore, likely to interact with plasma membranes and play a significant role in modulating cellular functions. So far, the details of the processes occurring at the interface between the extracellular matrix and cellular membranes are not fully understood.
View Article and Find Full Text PDFModeling Chromatography Binding through Molecular Dynamics Simulations with Resin Fragments.
J Phys Chem B
June 2024
In Silico Discovery, Janssen Research & Development, LLC, a Johnson & Johnson company, Spring House, Pennsylvania 19002, United States.
Accurate atomistic modeling of the interactions of a chromatography resin with a solute can inform the selection of purification conditions for a product, an important problem in the biotech and pharmaceutical industries. We present a molecular dynamics simulation-based approach for the qualitative prediction of interaction sites (specificity) and retention times (affinity) of a protein for a given chromatography resin. We mimicked the resin with an unrestrained ligand composed of the resin headgroup coupled with successively larger fragments of the agarose backbone.
View Article and Find Full Text PDFHistidine-based ionizable cationic surfactants: novel biodegradable agents for hydrophilic macromolecular drug delivery.
Drug Deliv Transl Res
September 2024
Department of Pharmaceutical Technology, Institute of Pharmacy, Center for Chemistry and Biomedicine, University of Innsbruck, Innsbruck, 6020, Austria.
The aim of this study was to design surfactants based on histidine (His) for hydrophobic ion-pairing and evaluate their safety and efficacy. Lauryl, palmitoyl and oleyl alcohol, as well as 2-hexyl-1-decanol were converted into surfactants with histidine as head-group via esterification. The synthesized His-surfactants were characterized regarding pK, critical micellar concentration (CMC), biodegradability, toxicity on Caco-2 cells, and ability to provide endosomal escape.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!