Interactions of Polydopamine Nanoparticles with Serine and Cysteine Proteases: Implications for Enhancing Protein Stabilization and Enzyme Activity

The interactions between polydopamine nanoparticles (PDA NPs) with serine protease α-chymotrypsin (α-CT) and cysteine protease stem bromelain (SB) have prompted significant emphasis due to their potential implications in biocatalysis and therapeutic applications. This study addresses the structural integrity, binding kinetics, and catalytic potential of α-CT and SB in the presence of PDA NPs by employing a combination of spectroscopic techniques and thermodynamic and kinetic analyses. The spectroscopic results reveal that PDA NPs bind to both enzymes (α-CT and SB) to create a stable complex without disrupting their active sites. Furthermore, fluorescence, circular dichroism (CD), and dynamic light scattering (DLS) analysis clearly demonstrate that the binding between α-CT and PDA NPs is strong compared to SB + PDA NPs. The thermodynamic data implies that PDA NPs significantly enhance the thermal stability of both α-CT and SB, as evidenced by the increased thermal transition temperature ( T m ) by 2.68 ± 0.50 °C in α-CT and 3.35 ± 0.50 °C in SB. Interestingly, ζ-potential and microscopic results such as TEM and FESEM pointed toward the adsorption of both proteases over the surface of PDA NPs. Additionally, spectrophotometric assay and kinetic studies confirm the retention of enzyme activity of both proteases upon conjugation with PDA NPs. Further, kinetic ( V max, K m ) and thermodynamic (Δ G u, Δ H m, Δ S m, ΔC p ) parameters also concluded the formation of a stable enzyme–NP complex. This study conveys the potential of PDA NPs in developing advanced enzyme–NPs integrated for enhancing protein stability and activity in diverse biomedical and industrial applications.