Conformational Differences in the Light Chain Constant Domain of Immunoglobulin G and Free Light Chain May Influence Proteolysis in AL Amyloidosis.

Immunoglobulin light chain amyloidosis (AL) is a life-threatening disease caused by the deposition of light chain (LC) and its fragments containing variable (V) and portions of constant (C) domains. AL patients feature either monoclonal free LCs (FLCs) circulating as covalent and noncovalent homodimers, or monoclonal immunoglobulin (Ig) wherein the LC and heavy chain (HC) form disulfide-linked heterodimers, or both. The role of full-length Ig in AL amyloidosis is unclear as prior studies focused on FLC or V domain. We used a mammalian cell-based expression system to generate four AL patient-derived full-length IgGs, two non-AL IgG controls, and six corresponding FLC proteins derived from an IGLV6-57 germline precursor. Comparison of proteins' secondary structure, thermal stability, proteolytic susceptibility, and disulfide link reduction suggested the importance of local vs. global conformational stability. Analysis of IgGs vs. corresponding FLCs using hydrogen-deuterium exchange mass spectrometry revealed major differences in the local conformation/dynamics of the C domain. In all IgGs vs. FLCs, segments containing β-strand and α-helix βA-αAB were more dynamic/exposed while segment βD-βE was less dynamic/exposed. Notably, these segments overlapped proteolysis-prone regions whose in vivo cleavage generates LC fragments found in AL deposits. Altogether, the results suggest that preferential cleavage in segments βA-αAB of FLC or βD-βE of LC in IgG helps generate amyloid protein precursors. We propose that protecting these segments using small-molecule stabilizers, which bind to the interfacial cavities C-C in FLC and/or C-C in IgG, is a potential therapeutic strategy to complement current approaches targeting V-V or V-C stabilization of LC dimer.