title: Carbohydrate recognition properties of human ficolins: glycan array screening reveals the sialic acid binding specificity of M-ficolin.
abstract: Ficolins are oligomeric innate immune recognition proteins consisting of a collagen-like region and a fibrinogen-like recognition domain that bind to pathogen- and apoptotic cell-associated molecular patterns. To investigate their carbohydrate binding specificities, serum-derived L-ficolin and recombinant H- and M-ficolins were fluorescently labeled, and their carbohydrate binding ability was analyzed by glycan array screening. L-ficolin preferentially recognized disulfated N-acetyllactosamine and tri- and tetrasaccharides containing terminal galactose or N-acetylglucosamine. Binding was sensitive to the position and orientation of the bond between N-acetyllactosamine and the adjacent carbohydrate. No significant binding of H-ficolin to any of the 377 glycans probed could be detected, providing further evidence for its poor lectin activity. M-ficolin bound preferentially to 9-O-acetylated 2-6-linked sialic acid derivatives and to various glycans containing sialic acid engaged in a 2-3 linkage. To further investigate the structural basis of sialic acid recognition by M-ficolin, point mutants were produced in which three residues of the fibrinogen domain were replaced by their counterparts in L-ficolin. Mutations G221F and A256V inhibited binding to the 9-O-acetylated sialic acid derivatives, whereas Y271F abolished interaction with all sialic acid-containing glycans. The crystal structure of the Y271F mutant fibrinogen domain was solved, showing that the mutation does not alter the structure of the ligand binding pocket. These analyses reveal novel ficolin ligands such as sulfated N-acetyllactosamine (L-ficolin) and gangliosides (M-ficolin) and provide precise insights into the sialic acid binding specificity of M-ficolin, emphasizing the essential role of Tyr(271) in this respect.
authors.affiliation: Laboratoire d'Enzymologie Moléculaire, Institut de Biologie Structurale Jean-Pierre Ebel, Commissariat à l'Energie Atomique, CNRS UMR 5075, Université Joseph Fourier, 41 rue Jules Horowitz, Grenoble 38027 Cedex 1, France.
authors.forename: Evelyne
authors.initials: E
authors.lastname: Gout
authors.forename: Virginie
authors.initials: V
authors.lastname: Garlatti
authors.forename: David F
authors.initials: DF
authors.lastname: Smith
authors.forename: Monique
authors.initials: M
authors.lastname: Lacroix
authors.forename: Chantal
authors.initials: C
authors.lastname: Dumestre-Pérard
authors.forename: Thomas
authors.initials: T
authors.lastname: Lunardi
authors.forename: Lydie
authors.initials: L
authors.lastname: Martin
authors.forename: Jean-Yves
authors.initials: JY
authors.lastname: Cesbron
authors.forename: Gérard J
authors.initials: GJ
authors.lastname: Arlaud
authors.forename: Christine
authors.initials: C
authors.lastname: Gaboriaud
authors.forename: Nicole M
authors.initials: NM
authors.lastname: Thielens
mesh_terms: D002241:Carbohydrates / Q000737:chemistry*; D018360:Crystallography, X-Ray; D005340:Fibrinogen / Q000737:chemistry / Q000235:genetics; D006801:Humans; D037102:Lectins / Q000737:chemistry / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D019158:N-Acetylneuraminic Acid / Q000737:chemistry / Q000378:metabolism*; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D040081:Protein Array Analysis; D011485:Protein Binding; D014443:Tyrosine; D000097604:Ficolins
chemical_list: D002241:Carbohydrates; D005340:Fibrinogen; D037102:Lectins; D019158:N-Acetylneuraminic Acid; D011134:Polysaccharides; D014443:Tyrosine; D000097604:Ficolins