Interheavy disulfide bridge in immunoglobulin G (IgG) reacting with dithionitrobenzoate: A unique feature in serum proteins

Abstract

Immunoglobulin G (IgG) of many species contains `labile' disulfide bonds (SS*), which within 24 h undergo a disulfide exchange with dithionitrobenzoic acid (NBSSBN). Aims of the present study were to detect directly this type of SS* groups by means of 14C-labelled NBSSBN, and to investigate its possible presence in other serum proteins. NBSSBN reacts during the first 30 min of incubation with free SH groups, and thereafter with the sulfur atoms of SS* groups. The latter reaction reaches equilibrium after 24 h. The total of thionitrobenzoate residues (NBS) bound to IgG is called `ΣS' and represents both SH and SS*. The results can be summarized as follows: (1) The measurement of the binding of 14C-NBSSBN gave identical ΣS values with IgG from humans and mice, as compared to the detection with the unlabelled reagent, which is based on the photometric determination of liberated NBS anions; whereas with IgG from rats some differences were observed which were ascribed to different batches of animals investigated; (2) experiments with electrophoretically separated serum proteins revealed only the γ-globulins strongly binding 14C-NBSSBN in addition to the 30 min reaction, which indicates that SS* is confined to the γ-globulin fraction; and (3) the significant decrease of ΣS in association with malignant tumors in man and animal models, which was previously described to be due to a specific alteration of the IgG subclass pattern, was likewise detected with the radiometrical method. Previous studies have identified SS* as one of the two inter-heavy disulfide bridges in IgG1, and possible implications of this group in specific functions of IgG1 are discussed.