title: Neutralizing monoclonal antibody epitopes of the Entamoeba histolytica galactose adhesin map to the cysteine-rich extracellular domain of the 170-kilodalton subunit.
abstract: Entamoeba histolytica adheres to human colonic mucins and colonic epithelial cells via a galactose-binding adhesin. The adhesin is a heterodimeric glycoprotein composed of 170- and 35-kDa subunits. Fragments of the hgl1 gene encoding the 170-kDa subunit were expressed as recombinant fusion proteins in Escherichia coli and reacted with anti-adhesin monoclonal antibodies (MAbs) or pooled human immune sera. The MAbs tested recognize seven distinct epitopes on the 170-kDa subunit and have distinct effects on the adherence and complement-inhibitory activities of the adhesin. All seven MAbs reacted with a fusion protein containing the cysteine-rich domain of the protein. Pooled human immune sera reacted with the same cysteine-rich domain as the MAbs and also with a construct containing the first 596 amino acids. Reactivity of three MAbs with the surface of intact trophozoites confirmed that the cysteine-rich domain was located extracellularly. The location of individual epitopes was fine mapped by constructing carboxy-terminal deletions in the cysteine-rich region of the fusion protein. The locations of adherence-enhancing and -inhibiting epitopes were partially distinguished, and the epitopes where complement-inhibitory MAbs bound were demonstrated to be near the adhesin's area of sequence identity with the human complement inhibitor CD59.
authors.affiliation: Department of Medicine, University of Virginia, Charlottesville 22908.
authors.forename: B J
authors.initials: BJ
authors.lastname: Mann
authors.forename: C Y
authors.initials: CY
authors.lastname: Chung
authors.forename: J M
authors.initials: JM
authors.lastname: Dodson
authors.forename: L S
authors.initials: LS
authors.lastname: Ashley
authors.forename: L L
authors.initials: LL
authors.lastname: Braga
authors.forename: T L
authors.initials: TL
authors.lastname: Snodgrass
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000913:Antibodies, Protozoan / Q000276:immunology; D000953:Antigens, Protozoan / Q000737:chemistry*; D000954:Antigens, Surface / Q000276:immunology*; D001483:Base Sequence; D003545:Cysteine; D004252:DNA Mutational Analysis; D019286:Endopeptidase K; D004748:Entamoeba histolytica / Q000276:immunology*; D000939:Epitopes; D005110:Extracellular Space / Q000276:immunology; D037161:Galectins; D006388:Hemagglutinins / Q000276:immunology*; D006801:Humans; D066298:In Vitro Techniques; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000276:immunology; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D010504:Periodic Acid / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D017384:Sequence Deletion; D012697:Serine Endopeptidases / Q000494:pharmacology
chemical_list: D000911:Antibodies, Monoclonal; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D000954:Antigens, Surface; D000939:Epitopes; D037161:Galectins; D006388:Hemagglutinins; D008562:Membrane Glycoproteins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D010504:Periodic Acid; D012697:Serine Endopeptidases; D019286:Endopeptidase K; D003545:Cysteine