title: Differential reduction of the inter-chain disulfide bonds of rat immunoglobulin E: relation to biological activity.
abstract: Monoclonal rat IgE was reduced over a range of dithiothreitol (DTT) concns. The number of disulfide bonds reduced and their location in the IgE molecule were studied. One millimolar DTT was found to split the two inter-heavy-chain disulfide bonds of the C epsilon 2 domain while increasing DTT concn to 10 mM split the two inter-heavy-light-chain disulfide bridges. Therefore, the sensitivities to reduction of disulfide bonds in rat IgE were found to be the opposite of those in human IgE. In addition, the results indicated the absence, in rat IgE, of the intra-epsilon-chain labile disulfide bond of the C epsilon 1 domain, which is reduced by 2 mM DTT in human IgE. Circular dichroism studies showed significant modifications, mainly of tertiary structure, for rat IgE reduced with 10 mM DTT, but not for IgE reduced with 1 mM DTT. The ability to block passive sensitization with reaginic antibody was not modified when IgE was reduced with 1 mM DTT (which split the two inter-heavy-chain disulfide bonds), but was lost when inter-heavy-light-chain bridges were reduced with 10 mM DTT. In addition, a non-covalent epsilon-chain dimer was found to have the same blocking activity as native IgE (or IgE reduced with 1 mM DTT). Therefore, the results suggest that reduction of most or all the inter-chain disulfide bonds, in rat as in human IgE, induces changes in quaternary structure, more especially in the relationship between the Fab and Fc parts of the molecule, leading to steric blockade, by Fab, of the binding sites for mast cells present on Fc.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D004220:Disulfides; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D007073:Immunoglobulin E / Q000276:immunology*; D007128:Immunoglobulin Fragments / Q000276:immunology; D010084:Oxidation-Reduction; D011487:Protein Conformation; D051381:Rats; D013329:Structure-Activity Relationship
chemical_list: D004220:Disulfides; D000939:Epitopes; D007128:Immunoglobulin Fragments; D007073:Immunoglobulin E; D004229:Dithiothreitol