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JBC, Vol. 250, Issue 10, 3891-3896, May, 1975
S. P. Hauptman and T. B. Tomasi Jr
Employing mercaptoethylamine as a reducing agent, it was demonstrated by
analytical ultracentrifugation and polyacrylamide gel electrophoresis that
polymeric immunoglobulin A (IgA) was reduced to a 10 S dimer and 7 S
monomer, and that dimer IgA was more resistant to reductive cleavage than
the higher polymers. When dimer and monomer IgA were subjected to
electrophoresis in polyacrylamide gels in 8 M urea or chromatographed on
Bio-Gel P-200 equilibrated in 4 M guanidine HCl, there was no dissociations
into H, L, or J chains, suggesting that the interchain disulfide bridges
between H--H, L--H, and H--J were intact and that mercaptoethylamine
produced selective cleavage of intersubunit bonds. Only the dimer, with a
sedimentation coefficient of 10.2 S, released J chain upon reduction with
dithiothreitol. Polymers of IgA were reduced with mercaptoethylamine and
subsequently alkylated with [14C]-iodoacetamide and the dimer and monomer
isolated. The results demonstrated that the isolated dimer contained 2 mol
of [14C]labeled S carboxyamidomethylcysteine per mol of dimer, while the
monomer contained 1 mol of --SH per mol of monomer. The labeled dimer was
then completely reduced with dithiothreitol and alkylated with
[14C]iodoacetamide and J chain isolated. It was shown that the J chain
contained no 14C-labeled sulfhydryl groups, while the monomer contained 1
mol of --SH per mol of monomer. These results suggest that J chain is
disulfide-bonded to only two of the subunits of polymeric IgA and that the
remaining subunits in the higher polymers are disulfide-bonded one to the
other. This is similar to the model previously suggested for 19 S
immunoglobulin M (IgM). The sulfhydryl data also suggests that polymeric
IgA may not be a covalently bonded circular structure as has been shown for
IgM. However, no conclusions can be made from this study regarding the
structure of pentameric IgA, since this species was present in very small
amounts in our polymer preparation.
Mechanism of immunoglobulin A polymerization
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