Copper ion-dependent amyloid formation by Beta 2-microglobulin

beta2- microglobulin (beta2m) is the 99 residue non-covalent light chain associated with the class I Major Histocompatibility Complex where it plays a role in proper folding and expression. In patients with end-stage renal disease who are treated by hemodialysis therapy, beta2m aggregates and deposits as amyloid plaques systemically throughout the joints. Since this discovery, nearly two decades of work has focused on understanding the mechanism by which this normally soluble protein is converted into insoluble aggregates.;beta2m is a specific Cu2+-binding protein, and exposure to Cu2+ is sufficient to induce aggregation under near-physiological conditions. This aggregation is slow, occurring on the week(s) timescale, but is preceded by rapid oligomerization over the course of hours. These oligomers require bound metal for stability but mature amyloid remains stable in the presence of metal chelate.;The overall goal of this project is to understand how the binding of Cu2+ permits self-association of beta2m. Specifically, I have used mutagenesis to trap, isolate, and crystallize a Cu2+-bound hexameric intermediate. The structure of this oligomer reveals the location of bound metal, the detailed intermolecular interfaces, and a vast set of conformational changes related to metal binding. Of particular interest, we see the cis-trans isomerization of a conserved proline which has been previously implicated in both the folding and aggregation pathways of beta2m. In addition, by studying the sensitivity to metal chelate as a function of time, we established that the transition from C2+ -dependent to Cu2+-independent states occurs within small oligomers before mature amyloid formation. These states require initial exposure to Cu2+ to form but no longer require bound metal for stability, establishing a transient role for Cu2+ in amyloid formation by beta2m.