JBC INTERFERin siRNA transfection reagent

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JBC, Vol. 250, Issue 10, 3891-3896, May, 1975

Mechanism of immunoglobulin A polymerization

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.
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V. Sorensen, I. B. Rasmussen, V. Sundvold, T. E. Michaelsen, and I. Sandlie
Structural requirements for incorporation of J chain into human IgM and IgA
Int. Immunol., January 1, 2000; 12(1): 19 - 27.
[Abstract] [Full Text] [PDF]




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