Pathogenic D76N Variant of β2-Microglobulin: Synergy of Diverse Effects in Both the Native and Amyloid States

Abstract

β2-microglobulin (β2m), the light chain of the MHC-I complex, is associated with dialysis-related amyloidosis (DRA). Recently, a hereditary systemic amyloidosis was discovered, caused by a naturally occurring D76N β2m variant, which showed a structure remarkably similar to the wild-type (WT) protein, albeit with decreased thermodynamic stability and increased amyloidogenicity. Here, we investigated the role of the D76N mutation in the amyloid formation of β2m by point mutations affecting the Asp76-Lys41 ion-pair of WT β2m and the charge cluster on Asp38. Using a variety of biophysical techniques, we investigated the conformational stability and partial unfolding of the native state of the variants, as well as their amyloidogenic propensity and the stability of amyloid fibrils under various conditions. Furthermore, we studied the intermolecular interactions of WT and mutant proteins with various binding partners that might have in vivo relevance. We found that, relative to WT β2m, the exceptional amyloidogenicity of the pathogenic D76N β2m variant is realized by the deleterious synergy of diverse effects of destabilized native structure, higher sensitivity to negatively charged amphiphilic molecules (e.g., lipids) and polyphosphate, more effective fibril nucleation, higher conformational stability of fibrils, and elevated affinity for extracellular components, including extracellular matrix proteins.

BeStSel: webserver for secondary structure and fold prediction for protein CD spectroscopy

“…β 2 -Microglobulin (β2m) is the light chain of the major histocompatibility complex I. Dissociating from the complex, the protein circulates in monomeric form in the blood and is associated with dialysis related amyloidosis in long-term haemodialysis patients. In 2012, a variant of the protein carrying a D76N point mutation was discovered causing a hereditary systemic amyloidosis with pathophysiology strikingly different from that of the wild-type protein ( 39 , 40 ). The first investigations found that the mutant exhibits a high-resolution structure almost identical to that of the wild-type protein (Figure 2D ) and its unique behavior can be discovered by using a rather complex methodology.…”

The effect of mutation on an aggregation-prone protein: An in vivo, in vitro, and in silico analysis

“…3 A ). Indeed, a recent study has highlighted the importance of this salt bridge for D76N-β 2 m stability and aggregation ( 47 ). Interestingly, two of the most rapidly aggregating variants (D76N- and D76Q-β 2 m) share an amide sidechain.…”

Immune Checkpoint Inhibitors in Cancer Therapy—How to Overcome Drug Resistance?

“…Leukocyte immunoglobulin-like receptor B (LILRB) is expressed in most immune cells, and in binding to the major ligand major histocompatibility complex I (MHCI), mediates the negative regulation of immune cell activation [ 50 , 51 ]. MHCI is a complex formed by the HLAα chain and β 2-microglobulin (β2 M) [ 52 ]. Anchored by glycosyl phosphatidylinositol, CD24 interacts with sialic acid, binding immunoglobulin-like agglutinin-10 (SIGlec-10) to β2 M, which is overexpressed in some tumor tissues and binds to LILRB1 on macrophages to inhibit phagocytosis, resulting in a loss of immune surveillance [ 53 , 54 ].…”