title: Screening Glycolipids Against Proteins in Vitro Using Picodiscs and Catch-and-Release Electrospray Ionization-Mass Spectrometry.
abstract: This work describes the application of the catch-and-release electrospray ionization-mass spectrometry (CaR-ESI-MS) assay, implemented using picodiscs (complexes comprised of saposin A and lipids, PDs), to screen mixtures of glycolipids (GLs) against water-soluble proteins to detect specific interactions. To demonstrate the reliability of the method, seven gangliosides (GM1, GM2, GM3, GD1a, GD1b, GD2, and GT1b) were incorporated, either individually or as a mixture, into PDs and screened against two lectins: the B subunit homopentamer of cholera toxin (CTB5) and a subfragment of toxin A from Clostridium difficile (TcdA-A2). The CaR-ESI-MS results revealed that CTB5 binds to six of the gangliosides (GM1, GM2, GM3, GD1a, GD1b, and GT1b), while TcdA-A2 binds to five of them (GM1, GM2, GM3, GD1a, and GT1b). These findings are consistent with the measured binding specificities of these proteins for ganglioside oligosaccharides. Screening mixtures of lipids extracted from porcine brain and a human epithelial cell line against CTB5 revealed binding to multiple GM1 isoforms as well as to fucosyl-GM1, which is a known ligand. Finally, a comparison of the present results with data obtained with the CaR-ESI-MS assay implemented using nanodiscs (NDs) revealed that the PDs exhibited similar or superior performance to NDs for protein-GL binding measurements.
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.
authors.forename: Jun
authors.initials: J
authors.lastname: Li
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.
authors.forename: Xuxin
authors.initials: X
authors.lastname: Fan
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.
authors.forename: Elena N
authors.initials: EN
authors.lastname: Kitova
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.
authors.forename: Chunxia
authors.initials: C
authors.lastname: Zou
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.
authors.forename: Christopher W
authors.initials: CW
authors.lastname: Cairo
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Biological Sciences, University of Calgary , Calgary, Alberta, Canada T2N 1N4.
authors.forename: Luiz
authors.initials: L
authors.lastname: Eugenio
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Biological Sciences, University of Calgary , Calgary, Alberta, Canada T2N 1N4.
authors.forename: Kenneth K S
authors.initials: KK
authors.lastname: Ng
authors.affiliation: Department of Biochemistry, University of Toronto , Toronto, Ontario, Canada M5S 1A8.
authors.forename: Zi Jian
authors.initials: ZJ
authors.lastname: Xiong
authors.affiliation: Department of Biochemistry, University of Toronto , Toronto, Ontario, Canada M5S 1A8.|Princess Margaret Cancer Centre, University Health Network , Toronto, Ontario, Canada M5G 1L7.
authors.forename: Gilbert G
authors.initials: GG
authors.lastname: Privé
authors.affiliation: Alberta Glycomics Centre, Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.|Department of Chemistry, University of Alberta , Edmonton, Alberta, Canada T6G 2G2.
authors.forename: John S
authors.initials: JS
authors.lastname: Klassen
mesh_terms: D001427:Bacterial Toxins / Q000032:analysis*; D002772:Cholera Toxin / Q000032:analysis*; D004768:Enterotoxins / Q000032:analysis*; D005732:Gangliosides / Q000737:chemistry*; D006801:Humans; D037102:Lectins / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D001427:Bacterial Toxins; D004768:Enterotoxins; D005732:Gangliosides; D037102:Lectins; C041348:tcdA protein, Clostridium difficile; D002772:Cholera Toxin