On-Membrane Renaturation of Recombinant Ro60 Autoantigen by Calcium Ions

Abstract

Calcium plays an important role in many biochemical processes in different cell types. This divalent cation interacts with specific calcium-binding proteins that serve as calcium sensors and regulatory proteins to mediate its function. Previously, we found that calcium was involved in the protein-protein interaction observed between Ro60 multiple antigenic peptides and Ro60 autoantigen. Since calcium bound Ro60 multiple antigenic peptides, we hypothesized that it would renature human recombinant Ro60 on a protein blot. We found that anti-Ro60 antibodies bound significantly higher to the recombinant Ro60 antigen that was incubated with calcium compared to that incubated without calcium on a polyvinylidene fluoride (PVDF) blot. Since the immunological epitopes of Ro60 are mainly conformational, we believe that calcium induced a more native tertiary structure in recombinant Ro60 autoantigen following blotting to a PVDF membrane.