A novel pentamer versus pentamer approach to generating neutralizers of verotoxin 1.
Verotoxins (VTs), or shiga-like toxins, are produced by enterohemorrhagic Escherichia coli (EHEC), which cause hemorrhagic colitis and hemolytic uremic syndrome. VTs are the major virulence factors in EHEC infection due to their cytotoxicity to various types of cells. Here, we present a novel type of VT neutralizer based on pentavalent single-domain antibodies, or pentabodies. Two single-domain antibodies (sdAbs) specific for the receptor binding sites of the B subunit of VT1 (VT1B) were isolated from a naïve llama phage display library. These two sdAbs were pentamerized to generate pentameric VT neutralizers, VTI-1 and VTI-3. Both VT neutralizers bound wild type VT1B specifically with superior functional affinity. In vitro neutralization assays showed that VTI-1 and VTI-3 were able to neutralize 90% and 40%, respectively, of the cytotoxicity caused by VT1. This effort provides the basis of a novel type of VT neutralizer that can potentially be produced at a relatively low cost.
Stone
Emily
E
Institute for Biological Sciences, National Research Council of Canada, 100 Sussex Dr., Ottawa, Ont. K1A 0R6, Canada.
Hirama
Tomoko
T
Chen
Wangxue
W
Soltyk
Anna L
AL
Brunton
James
J
MacKenzie
C Roger
CR
Zhang
Jianbing
J
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification; D000915:Antibody Affinity / Q000276:immunology; D001665:Binding Sites; D002161:Camelids, New World / Q000276:immunology; D002522:Chlorocebus aethiops; D003602:Cytotoxicity, Immunologic; D007700:Kinetics; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000276:immunology; D009500:Neutralization Tests; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011956:Receptors, Cell Surface / Q000276:immunology; D022622:Shiga Toxin 1 / Q000037:antagonists & inhibitors / Q000276:immunology*; D014709:Vero Cells
D000906:Antibodies; D050505:Mutant Proteins; D011956:Receptors, Cell Surface; D022622:Shiga Toxin 1