Peroxidase-like Catalytic Activity of Copper-Mediated Protein-Inorganic Hybrid Nanoflowers and Nanofibers of β-Lactoglobulin and α-Lactalbumin: Synthesis, Spectral Characterization, Microscopic Features, and Catalytic Activity

Abstract

A free Cys-SH-containing protein, β-lactoglobulin (β-LG), and another protein not possessing the same, viz., apo-α-lactoglobulin (apo-α-LA), were used in studies to demonstrate the role of this amino acid, along with its secondary structure, in the formation of a protein dimer and a protein-inorganic hybrid nanoflower and in the creation of the peroxidase-like activity of the nanomaterials produced when the proteins were treated with varying Cu2+ concentration under different pH conditions. An increase in the pH as well as the Cu2+ mole ratio results in increasing dimer formation in case of β-LG due to the presence of free Cys121-SH, while the dimer is not formed in case of apo-α-LA under the same conditions. The role of Cys in the dimer formation has been demonstrated both by MALDI and sodium dodecyl sulfate-polyacrylamide gel electrophoresis studies. Both of the proteins exhibited changes in their secondary structures to different extents as a function of pH, and the structures were stabilized by Cu2+ interactions, as studied by CD and fluorescence spectroscopy. The small and spherical nanoparticles formed at pH 7 with lower equivalents of Cu2+ join together to form larger aggregates at higher equivalents of Cu2+. For the same concentration at pH 9, both the aggregates and the nanoflowers were noticed. However, at pH 12, the Cu2+ binding induces the formation of fibers along with the flowers. Both the nanoflowers and nanofibers exhibited peroxidase-like activity in a catalytic manner. Nanoflowers were also shown to detect phenol in the concentration range from 10 to 200 μM. The copper-induced nanobiomaterial obtained in the case of apo-α-LA also exhibited peroxidase-like activity. Thus, this paper deals with the green synthesis of copper-induced protein (β-LG/apo-α-LA)-inorganic hybrid nanomaterials that are important due to their applications as nanobiomaterials.