IgA Subclasses Research Articles

A Study of the Association of Human Secretory Component with IgA and IgM Proteins

Human secretory component (SC) was isolated from colostral whey, and the binding of 125I-SC to purified IgA and IgM monoclonal proteins was studied using two methods to separate free from immunoglobulin-bound 125I-SC: a) gel filtration on Sephadex G-200, and b) precipitation of 125I-SC-Ig complexes with anti-Ig antibody. Both IgA dimeric proteins and IgM pentamers bound 125I-SC with approximately one SC-binding site per mole of polymer and similar affinity. Assuming a reversible equilibrium, an apparent association constant congruent to 10-8 M-1 was calculated to govern the binding of 125I-SC to immunoglobulin polymers. The assignment of a single association constant may be an oversimplication, particularly for the case of IgA polymers, since evidence was obtained that disulfide bonds were formed in the 125I-SC-IgA complex. Despite the complexity of the reaction, binding of 125I-SC to both IgA and IgM polymers could be analyzed by standard methods of saturation analysis, and both were shown to have a similar affinity for 125I-SC. No differences were noted in the affinity of 125I-SC binding to the IgA1 and IgA2 subclasses. Binding of monomeric IgA and IgM proteins could not be measured and was at least 100-fold lower than that found for IgA and IgM polymers. Complexes of 125I-SC with IgA dimers were presumed to involve covalent bond formation, since these complexes did not dissociate in guanidine-HCl. One IgA2 trimer did not form a covalent bond since it was completely dissociated in guanidine. In contrast, 125I-SC-IgM complexes were dissociated in denaturing solvent, indicating that such complexes were held together primarily by non-covalent bonds. Experiments with (Fc)5 mu isolated by high temperature tryptic digestion of IgM showed that binding of 125I-SC was to the Fc region of IgM proteins. It was suggested that the binding of SC with similar affinity to both IgA and IgM polymers may be important in the biologic function of both these immunoglobulin classes.

Partial amino acid sequence of an IgA2 human immunoglobulin heavy chain

The amino acid sequence of the heavy chain of an IgA2, A2m(1) polymeric myeloma protein (Avi) was studied. Altogether, sequence data were obtained for some 130 residues. Including the amino acids placed by homology with IgA 1, this accounts for some 170 residues, thus representing more than one-third of the α 2 chain. The sequence includes 26 amino acids from the amino-terminal end (V H III), and 25 residues at the “hinge” region. Of a total of 17 cysteine residues, 14 were located in regions of the molecule which were identical or homologous in the α 1 and α 2 chains. These striking homologies, together with the results obtained by diagonal maps of the hinge region, suggest a similar arrangement for the disulfide bridges in both subclasses of IgA. Study of the cysteine-containing peptides of the J chain are consistent with the conclusion that the J chains associated with different classes of immunoglobulins are identical.

Detection of human IgA subclasses and allotypes by polymer-induced precipitation in agarose gel

Only some human sera containing anti-IgA antibodies have precipitating properties, so that usually the presence of these isoantibodies is established by hemagglutination and their specificity by hemagglutination inhibition. The incorporation of a high molecular weight polymer, polyethylene glycol (PEG), will convert the system to a precipitating one in agarose gel. IgA subclass specificities are readily demonstrated in the presence of PEG alone. The addition of isolated human Clq to the system will enhance the precipitation of otherwise soluble small molecular weight complexes so that the A 2m allotype specificities can be seen as well. This simplified procedure facilitates the detection of these clinically significant isoantibodies in patients with IgA deficiency.

Production of α1 and α2 Immunoglobulin Heavy Chains during Fetal Life

Summary IgA and its subclasses IgA1 and IgA2 were measured in fetal serum and amniotic fluid throughout gestation by hemagglutination inhibition tests. The IgA1/(IgA1 + IgA2) ratio was higher for fetal serum than for amniotic fluid and was significantly higher in the third than in the second trimester in both amniotic fluid and fetal serum. Am2, a genetic marker for human α2 heavy chains, was typed in a number of combinations of maternal and paternal sera, amniotic fluid, and fetal serum. Three families were informative in that Am2 was present in amniotic fluid, was absent from maternal serum, but present in paternal serum. It was concluded that the fetus synthesized IgA2 in vtero.

Special characteristics of the IgA2 subclass.

Although in man IgA is a relatively minor fraction of circulating serum immunoglobulins, lately it has been the object of a burgeoning research effort. Mainly this is due to the realization that IgA is the principal immunoglobulin in external secretions, where it is the prime mediator of local humoral immunity on mucous membrane surfaces. In addition, the genetics of human IgA has been advanced by the discovery of genetic markers on the IgA2 subclass. The Am2 marker is linked to and behaves like the Gm markers of IgG which have been so thoroughly studied. The other marker involves the presence or absence of the light-to-heavy interchain disulfide bridges in IgA2 proteins. The absence of these bridges in one of the genetic variants is associated with unusual structural properties.

Distribution of IgA subclasses in sera and bone marrow plasma cells of 21 normal individuals.

I would like to report some experiments which Dr. Morell and I have recently performed in Berne, Switzerland. We compared the distribution of the two IgA subclasses in the sera and in the IgA containing cells in the bone marrow of normal individuals. We are indebted to Dr. Hijmans for his help in the immunofluorescence studies of the bone marrow.

The carbohydrate composition of glycopeptides isolated from IgA2 immunoglobulin.

Analyses of the carbohydrate content of eleven IgA1 and six IgA2 myeloma proteins revealed that, besides the known restriction of galactosamine to IgA1 subclass (1,2), statistically significant quantitative differences exist between the carbohydrate composition of the two subclasses. The greater amounts of fucose, mannose, and galactosamine in IgA2 immunoglobulins explain the higher total carbohydrate content of these proteins (Table 1). No statistical differences were found in the galactose and sialic acid content of the subclasses. The relatively high content of fucose in IgA2 immunoglobulins suggests that these proteins contain more highly branched oligosaccharides than proteins of the IgA1 subclass.KeywordsSialic AcidAmmonium BicarbonateCarbohydrate CompositionSialic Acid ContentAmmonium Bicarbonate BufferThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

J-chain determinants in polymeric immunoglobulins.

AbstractAn antiserum was raised against purified J‐chain from S‐sulfonated human secretory IgA and was tested for its reactivity with secretory IgA, polymeric myeloma proteins of IgA1 and IgA2 subclasses and a Waldenström IgM macroglobulin. The antiserum, which was absorbed with normal human IgG and L‐chains, developed a single precipitin line with the following S‐sulfonated proteins: purified J‐chain, secretory IgA, polymeric IgA1, and IgA2, and IgM. The antiserum also reacted with the same proteins after complete reduction and alkylation. The antiserum did not precipitate the same polymeric immunoglobulins in their native states. At high concentrations, however, native polymeric serum IgA and IgM inhibited the precipitin reaction between the purified J‐chain and its antibodies. In contrast, native secretory IgA displayed no inhibition in this system. After treatment of these immunoglobulins with 8 M urea, 5 M guanidine, or 1 M propionic acid, the J‐chain determinants became accessible and were precipitated with the anti‐J‐chain antiserum, with the exception of IgM which displayed very weak reactivity only after exposure to 5 M guanidine. In conclusion, J‐chain determinants are accessible only to a very limited extent in native serum polymeric IgA and IgM, and none are detectable in the native secretory IgA. Most of the determinants become easily available by mere denaturation of these immunoglobulins with exposure to several kinds of dissociating agents.

The Release of Granule Enzymes from Human Neutrophils Stimulated by Aggregated Immunoglobulins of Different Classes and Subclasses

Abstract Aggregated human myeloma proteins of subclasses IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2 and normal IgG have been shown to react with human neutrophils in a serum-free medium and to induce release of the lysosomal enzyme β glucuronidase to the exterior of the cells. In contrast, IgD, IgE and IgM macroglobulins did not stimulate this release. Incubation of neutrophils with insoluble IgG and IgA aggregates resulted in adherence of the aggregates to the neutrophils and their phagocytosis. During this process, release of granule enzymes, but not of the cytoplasmic enzyme lactic dehydrogenase, occurred, indicating a selective process rather than one of cell lysis. Soluble aggregates did not induce liberation of enzymes when incubated with the neutrophils in suspension, but when bound to micropore filters to serve as nonphagocytosable surfaces, they stimulated adherent cells to release their granule contents. This latter process has previously been shown to result from direct extrusion of neutrophil granules to the exterior.

Close Linkage of the Genes for Rabbit IgA Allotypes, Af71 to Af75, to the Chromosomal Region Controlling Immunoglobulin Heavy Chain Allotypes

Abstract Five allotypes of rabbit IgA immunoglobulin, designated Af71, Af72, Af73, Af74 and Af75, have been identified in this laboratory and found to be inherited in linkage groups of f71 and f75, f72 and f74 or f73 and f74 as if they were controlled by three autosomal codominant alleles f71,75, f72,74 and f73,74, at a chromosomal region designated the f locus (1, 2).3 Tentatively, the f locus or region can be thought of as two linked regions or subloci such that one set of alleles controls f71, f72 and f73 and another set, controls f74 and f75. These five allotypic specificities are not found on IgG or IgM and thus appear to be associated uniquely with serum and secretory IgA (1). Furthermore, the phenogroup pair of allotypic specificities do not appear to be on the same IgA molecule but on separate IgA molecules, perhaps on subclasses of IgA (3).

WOOLHOUSE (1666-1733-4)

OBJECTIVE--To study the effects of sulphasalazine (SASP) on the systemic and mucosal humoral immune systems in patients with rheumatoid arthritis (RA). METHODS--Serum concentrations of interleukin 6 (IL-6), class and subclass specific IgG, IgA and IgM, IgA and IgG antigliadin antibodies and rheumatoid factors (RF) of IgG, IgA (including IgA1 and IgA2 subclasses) and IgM isotypes were measured before and 16 weeks after sulphasalazine (SASP) therapy in 15 female and three male patients with RA. Amounts of immunoglobulins in saliva and jejunal fluid were measured as estimates of mucosal humoral immunity. RESULTS--Serum concentrations of IgA and IgG decreased significantly during SASP therapy and correlated with reduced concentrations of IL-6. In addition, levels of circulating IgA RF, IgA anti-gliadin antibodies and IgM RF decreased significantly after the treatment. In contrast, immunoglobulin levels in saliva and jejunal fluid were unaltered. CONCLUSION--SASP exerts powerful but selective inhibitory effects on systemic immunoglobulin production, whereas no effects on mucosal immunoglobulin production were observed. The decreased systemic B cell activity may be mediated by downregulation of the production of IL-6, a cytokine with Ig switching properties.