Binding and multiple hydrolytic sites in epitopes recognized by catalytic anti-peptide antibodies.
Autoantibodies purified from humans catalyse the hydrolysis of the neurotransmitter, vasoactive intestinal peptide (VIP). Evidence that the hydrolysis of VIP is due to antibodies includes: the antibody preparations are free of detectable non-immunoglobulin (non-Ig) contamination; the hydrolytic activity is removed by precipitation with anti-human IgG antibody; human B lymphoblastoid cells transformed with Epstein-Barr virus secrete hydrolytic antibodies in culture; the Fab fragments of the antibodies exhibit VIP hydrolysis; and affinity chromatography on immobilized VIP permits purification of specific antibodies with greatly enriched hydrolytic and binding activities. One of the catalytic antibody preparations hydrolyses the Gln-16-Met-17 bond. Studies with synthetic VIP fragments showed that the epitope recognized by this antibody is formed by VIP(15-28). Important binding interactions are contributed by VIP(22-28), a sequence four residues distant from the scissile bond. Antibodies from a second subject hydrolyse six peptide bonds in VIP, clustered between residues 14 and 22. These bonds link amino acids of different charge, size and hydrophobicity, suggesting that the hydrolytic repertoire of the antibodies is considerable. The antibodies do not hydrolyse peptides unrelated in sequence to VIP. Cleavage of several peptide bonds in VIP by polyclonal antibody preparations may be due to several antibodies, each with a unique cleavage specificity. Alternatively, a single antibody may make catalytically productive contact at multiple peptide bonds in the substrate, because of conformational flexibility of VIP or of the antibody active site. Purified light chains from the catalytic antibodies hydrolysed VIP more rapidly than did intact antibodies. The residues constituting the catalytic site of an antibody may be encoded in germline V-region genes or may arise during maturation of the antibody response.
Paul
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S
Department of Pharmacology, University of Nebraska Medical Center, Omaha, 68198.
Johnson
D R
DR
Massey
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D000595:Amino Acid Sequence; D000906:Antibodies / Q000378:metabolism*; D000937:Antigen-Antibody Reactions / Q000276:immunology*; D001666:Binding Sites, Antibody / Q000276:immunology*; D002384:Catalysis; D000939:Epitopes / Q000276:immunology; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology*; D014660:Vasoactive Intestinal Peptide / Q000276:immunology
D000906:Antibodies; D000939:Epitopes; D010455:Peptides; D014660:Vasoactive Intestinal Peptide