Role of domain interactions in the aggregation of full-length immunoglobulin light chains

Amyloid light-chain (LC) amyloidosis is a protein misfolding disease in which the aggregation of an overexpressed antibody LC from a clonal plasma cell leads to organ toxicity and patient death if left untreated. While the overall dimeric architecture of LC molecules is established, with each LC composed of variable (V-L) and constant (CL) domains, the relative contributions of LC domain-domain interfaces and intrinsic domain stabilities to protection against LC aggregation are not well understood. To address these topics we have engineered a number of domain-destabilized LC mutants and used solution NMR spectroscopy to characterize their structural properties and intrinsic stabilities. Moreover, we used fluorescence spectroscopy to assay their aggregation propensities. Our results point to the importance of both dimerization strength and intrinsic monomer stability in stabilizing V-L domains against aggregation. Notably, in all cases considered V-L domains aggregate at least 10-fold faster than full-length LCs, establishing the important protective role of C-L domains. A strong protective coupling is found between V-L-V-L and C-L-C-L dimer interfaces, with destabilization of one interface adversely affecting the stability of the other. Fibril formation is observed when either the V-L or C-L domain in the full-length protein is severely destabilized (i.e., where domain unfolding free energies are less than 2 kcal/mol). The important role of C-L domains in preventing aggregation highlights the potential of the C-L-C-L interface as a target for the development of drugs to stabilize the dimeric LC structure and hence prevent LC amyloidosis.