Deciphering the Role of the Concentration and Reduction of Graphene Oxide Nanosheets in Their Interaction and Protein Corona (PC) Formation with Bromelain Proteins: A Spectroscopic and Biophysical Approach

Abstract Graphene oxide (GO) and its reduced form (RGO) constitute a novel class of applied materials that have recently gained global attention in various fields, particularly biomedicine, due to their excellent properties. To apply this class of nanomaterials in biomedical fields, a thorough understanding of their interaction with biomacromolecules is essential. Given the therapeutic importance of bromelain (BM) proteins, here, the impact of GO concentration and reduction on their interaction with bromelain has been examined using a variety of spectroscopic and biophysical approaches. The results indicate that the adsorption of BM proteins on the surface of GO/RGO occurs spontaneously and rapidly, forming a protein corona (PC). The binding affinity of BM with GO is much higher than with its reduced form (RGO). The binding of BM with GO occurs through cooperative interactions, whereas both cooperative and anticooperative binding are observed in the BM/RGO system. The adsorption of BM on GO nanosheets is primarily driven by strong electrostatic and H-bonding interactions, whereas weak hydrophobic interactions are the primary driving forces behind BM adsorption on the RGO surface. Distinct kinds of corona were observed in the GO/RGO-BM system based on the types of interactions involved (such as electrostatic, hydrophobic, H-bonding) between the protein and nanoparticles. BM exhibited a stronger and more stable association with GO, showing hard-corona-like characteristics, whereas its association with RGO was comparatively weaker and less stable, showing soft-corona-like characteristics. The therapeutic utility of this protein–nanoparticle system will be heavily dependent on its PC formation, which will be the focus of future research. This study lays the groundwork for future PC investigations on the diverse range of protein nanoparticles utilized in nanomedicine and environmental applications.