Probing the topography of free and polymeric Ig-bound human secretory component with monoclonal antibodies.
Secretory component (SC), an integral membrane protein expressed on basolateral surfaces of secretory epithelial cells, mediates the transport of polymeric Ig (PIg) into external secretions. The ectoplasmic segment of SC is released into secretions either in a free form (FSC) or bound to PIg as secretory IgA or IgM. The topography of human SC in its free and PIgA-bound form was studied by using mAb directed against each form of SC. Competition experiments identified a minimum of nine SC epitopes, one of which was dependent on an N-glycosidic moiety. Three of the polypeptide-derived epitopes were displayed on denatured, reduced, and alkylated SC, whereas the others were fully or partially dependent on the native conformation of SC. Epitopes recognized by the latter class of antibodies were mapped to discrete domains of SC, based on amino acid sequence and antibody-binding analysis of limited proteolytic fragments. One of the mAb (6G11), which was directed against an epitope on domain I of SC, inhibited the binding of FSC to PIgA. Overall, our results provide evidence that a region within domain I, as well as protease-sensitive interdomain regions of FSC, become masked or altered when SC binds to PIgA. Furthermore, the binding of SC to PIgA results in conformational changes, or formation of combinatorial epitopes, involving regions within domains II and III of SC but not domain V.
Department of Pediatrics, University of Texas Medical Branch, Galveston 77550.
M A
MA
Bakos
A
A
Kurosky
C S
CS
Woodard
R M
RM
Denney
R M
RM
Goldblum
D000478:Alkylation; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001667:Binding, Competitive; D015153:Blotting, Western; D000939:Epitopes; D006801:Humans; D007070:Immunoglobulin A / Q000276:immunology*; D007071:Immunoglobulin A, Secretory / Q000276:immunology; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000276:immunology; D011487:Protein Conformation; D012635:Secretory Component / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007070:Immunoglobulin A; D007071:Immunoglobulin A, Secretory; D046911:Macromolecular Substances; D010446:Peptide Fragments; D012635:Secretory Component; C034639:polymeric IgA