Adsorption of human carbonic anhydrase II variants to silica nanoparticlesoccur stepwise: Binding is followed by successive conformational changes to a molten-globule-like state

The surface adsorption behavior of protein variants of the enzyme human car bonic anhydrase II (HCA II) to silica nanoparticles has been investigated. Various destabilized mutants were produced by site-directed mutagenesis of amino acids located in the interior of the protein. The silica particles in duced a molten-globule-like state in all of the variants. All protein varia nts initially adsorbed to the particles, and then underwent conformational rearrangements in a stepwise manner, as indicated by the loss of activity a nd the subsequent loss of tertiary structure. Activity, CD, and ANS fluores cence measurements showed that a decrease in the global stability of the pr otein is strongly correlated to increased rates of conformational change fo llowing particle adsorption. In contrast to unfolding processes induced by chemical denaturants or heat, in the transition to the molten-globule-like state induced by the silica particles, the active site region unfolds befor e the majority of the tertiary interactions are broken.