ASSEMBLY OF IGM - ROLE OF DISULFIDE BONDING AND NONCOVALENT INTERACTIONS

Polymeric IgM is usually envisaged as an array of mu(2)L(2) monomers in which the mu heavy chains are held together by disulfide bonds involving cysteines at positions 337, 414, and 575. We have studied the importance of inter-mu-chain disulfide bonds for formation of IgM polymers and monomers by analyzing the effects of eliminating one or more of these disulfide bondings. Ablation of all inter-chain bonds by either chemical reduction and alkylation or by mutagenesis resulted in the exclusive production of halfmers (mu L) molecules. IgM composed of mu-chains bearing each of the other six possible combinations of cysteine to serine replacements was produced as different mixtures of polymers, monomers, and halfmers. Cysteine 575 was both necessary and sufficient for efficient assembly of IgM polymers and sufficient but not necessary for efficient assembly of monomers. Cysteine 337 was sufficient but not necessary for efficient assembly of monomers, and was neither sufficient nor necessary for formation of polymers. Cysteine 414 was neither necessary nor sufficient for efficient formation of either monomers or polymers. Data also suggest that noncovalent interactions between C mu 2 domains are stronger than the interactions between C mu 4/tail domains.