title: Mining the "glycocode"--exploring the spatial distribution of glycans in gastrointestinal mucin using force spectroscopy.
abstract: Mucins are the main components of the gastrointestinal mucus layer. Mucin glycosylation is critical to most intermolecular and intercellular interactions. However, due to the highly complex and heterogeneous mucin glycan structures, the encoded biological information remains largely encrypted. Here we have developed a methodology based on force spectroscopy to identify biologically accessible glycoepitopes in purified porcine gastric mucin (pPGM) and purified porcine jejunal mucin (pPJM). The binding specificity of lectins Ricinus communis agglutinin I (RCA), peanut (Arachis hypogaea) agglutinin (PNA), Maackia amurensis lectin II (MALII), and Ulex europaeus agglutinin I (UEA) was utilized in force spectroscopy measurements to quantify the affinity and spatial distribution of their cognate sugars at the molecular scale. Binding energy of 4, 1.6, and 26 aJ was determined on pPGM for RCA, PNA, and UEA. Binding was abolished by competition with free ligands, demonstrating the validity of the affinity data. The distributions of the nearest binding site separations estimated the number of binding sites in a 200-nm mucin segment to be 4 for RCA, PNA, and UEA, and 1.8 for MALII. Binding site separations were affected by partial defucosylation of pPGM. Furthermore, we showed that this new approach can resolve differences between gastric and jejunum mucins.
authors.affiliation: Institute of Food Research, Norwich Research Park, Norwich, UK.
authors.forename: A Patrick
authors.initials: AP
authors.lastname: Gunning
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Kirby
authors.forename: Christine
authors.initials: C
authors.lastname: Fuell
authors.forename: Carmen
authors.initials: C
authors.lastname: Pin
authors.forename: Louise E
authors.initials: LE
authors.lastname: Tailford
authors.forename: Nathalie
authors.initials: N
authors.lastname: Juge
mesh_terms: D000818:Animals; D005752:Gastric Mucins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D005753:Gastric Mucosa / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism; D037102:Lectins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D018625:Microscopy, Atomic Force / Q000379:methods; D009077:Mucins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D013057:Spectrum Analysis / Q000379:methods; D013552:Swine; D014018:Tissue Distribution
chemical_list: D005752:Gastric Mucins; D037102:Lectins; D009077:Mucins; D011134:Polysaccharides
keywords: adhesion; atomic force microscopy; lectin-carbohydrate interaction; molecular recognition