Expression, purification and biochemical characterization of recombinant murine secretory component: a novel tool in mucosal immunology.
Reconstitution of secretory IgA (S-IgA) by the association in vitro of secretory component (SC) and polymeric IgA (pIgA) obtained from hybridomas is a valuable tool in the study of the structure-function relationship in this particular class of antibody. Although dimeric IgA (dIgA) can be obtained and purified from hybridoma clones, SC remains tedious to isolate in sufficient amounts from colostral milk. Several murine models for the study of mucosal immunity are available, which could potentially benefit from the use of cognate IgA antibodies in various molecular forms, including dIgA and S-IgA. We report here on the establishment of two expression systems allowing the production of milligram amounts of pure recombinant murine SC (rmSC) with preserved murine pIgA-binding capability. The first system relies on the use of recombinant vaccinia virus to prompt infected HeLa cells to express the murine SC protein, whereas the second system is based on a stably transfected cell clone exhibiting murine glycosylation. The second source of rmSC will permit the study of the role of its sugar moieties in pathogen-host interactions, and the evaluation of its function in passive protection without risking adverse immune responses. The extensive biochemical characterization conducted in this study demonstrates that rmSC is a dependable and convenient alternative to the natural product, and indicates that the J chain is dispensable in the recognition of pIgA and SC in vitro, whereas it is required for proper pIgA-polymeric Ig receptor interaction in vivo.
Institut Suisse de Recherches Expérimentales sur le Cancer, CH-1066 Epalinges, Switzerland.
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Crottet
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Cottet
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Corthésy
D000818:Animals; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D018928:Immunity, Mucosal; D007070:Immunoglobulin A / Q000235:genetics / Q000276:immunology; D051379:Mice; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D012635:Secretory Component / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
D007070:Immunoglobulin A; D011994:Recombinant Proteins; D012635:Secretory Component