A conformational fingerprint for amyloidogenic light chains

Abstract Immunoglobulin light chain amyloidosis (AL) and multiple myeloma (MM) both share the overproduction of a clonal light chain (LC). However, while LCs in MM remain soluble in circulation, AL LCs misfold into toxic soluble species and amyloid fibrils that accumulate in organs, leading to distinct clinical manifestations. The significant sequence variability of LCs has hindered understanding of the mechanisms driving LC aggregation. Nevertheless, emerging biochemical properties, including dimer stability, conformational dynamics, and proteolysis susceptibility, distinguish AL LCs from those in MM under native conditions. This study aimed to identify a conformational fingerprint distinguishing AL from MM LCs. Using small-angle X-ray scattering (SAXS) under native conditions, we analyzed four AL and two MM LCs. We observed that AL LCs exhibited a slightly larger radius of gyration and greater deviations from X-ray crystallography-determined or predicted structures, reflecting enhanced conformational dynamics. SAXS data, integrated with molecular dynamics (MD) simulations, revealed a conformational ensemble where LCs adopt multiple states, with variable and constant domains either bent or straight. AL LCs displayed a distinct, low-populated, straight conformation (termed H state), which maximized solvent accessibility at the interface between constant and variable domains. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) experimentally validated this H state. These findings reconcile diverse experimental observations and provide a precise structural target for future drug design efforts. Competing Interest Statement The authors have declared no competing interest. Footnotes ↵* Stefano Ricagno, Email: stefano.ricagno{at}unimi.it In this version we: - moved one figure and one table from SI to main text to improve the presentation of SAXS/MD comparison (new figure 2 and new table 2) - reworked Figure 5 to improve the presentation of HDX-MS data - added three figures in SI to improve the analysis of SAXS data (new figure S1) and to provide evidence about the dimeric nature of LC (Figure S20, SEC-MALS; and Figure S21, SEC-SAXS) - moved all materials and methods in main text - improved abstract, introduction and discussion for readibility and to include additional literature evidence in support of our findings