d-Ribose glycates β2-microglobulin to form aggregates with high cytotoxicity through a ROS-mediated pathway

β2-Microglobulin (β2M) modified with advanced glycation end products (AGEs) is a major component of the amyloid deposits in hemodialysis-associated amyloidosis (HAA). However, the effect of glycation on the misfolding and aggregation of β2M has not been studied so far. Here we examine the molecular mechanism of aggregate formation of HAA-related ribosylated β2M in vitro. We find that the glycating agent d-ribose interacts with human β2M to generate AGEs that form aggregates in a time-dependent manner. Ribosylated β2M molecules are highly oligomerized compared with unglycated β2M, and have granular morphology. Furthermore, such ribosylated β2M aggregates show significant cytotoxicity to both human SH-SY5Y neuroblastoma and human foreskin fibroblast FS2 cells and induce intracellular reactive oxygen species (ROS). Presence of the antioxidant N-acetylcysteine (1.0 mM) attenuated intracellular ROS and prevented cell death induction in both SH-SY5Y and FS2 cells, indicating that the cytotoxicity of ribosylated β2M aggregates depends on a ROS-mediated pathway in both cell lines. In other words, d-ribose reacts with β2M and induces the ribosylated protein to form granular aggregates with high cytotoxicity through a ROS-mediated pathway. These findings suggest that ribosylated β2M aggregates could contribute to the dysfunction and death of cells and could play an important role in the pathogenesis of β2M-associated diseases such as HAA.

► We examine aggregation of disease-causing ribosylated β2-microglobulin. ► d-Ribose glycates β2-microglobulin to form ribosylated aggregates rapidly. ► Such ribosylated aggregates are highly oligomerized and have a granular morphology. ► Ribosylated aggregates show significant cytotoxicity to SH-SY5Y cells and induce ROS. ► Cytotoxicity of these ribosylated aggregates depends on a ROS-mediated pathway.