Impact of sonication on slurry shear -thinning of protein from sea cucumber (Apostichopus japonicus): Proteolytic reaction kinetics, thermodynamics, and conformational modification

The present study investigated the effect of slit dual-frequency ultrasonic (SDFU)-shear thinning on the kinetics and thermodynamic mechanism of sea cucumber protein (SCP) substrate enzymatic proteolysis reaction. Acoustic cavitation and mechanical shear force applied reduced the protein particle size, intermolecular forces, surface charge and increased the average kinetic energies of the molecules which resulted in substrate viscosity reduction. The enzyme-driven bonds were thus exposed for easy protease accessibility to enhance the reaction rate (k) and reduce Gibbs-free ( increment G) and activation energy (Ea) required. Substrate viscosity had a negative significant correlation (r = -0.865, p < 0.01) with SCP conversion degree. SDFU-pretreatment significantly boosted the reaction rate (ku) of enzymolysis at 20, 30, 40 and 50 degrees C by 63.0, 71.24, 37.5 and 37.22% respectively over the control (kc). Energy cost required for substrate conversion into product could be tremendously minimized by SDFU-pretreatment as affirmed by the lower values of enthalpy ( increment H), entropy ( increment S), and activation energy (Ea) (28.30 +/- 0.63kJmol- 1, -205.70 +/- 1.72 Jmol-1 k-1 and 31.01 +/- 0.27 kJmol-1 correspondingly) in comparison to the control. Higher cavitation yield and shear force from dual-frequency ultrasonication (20/68 kHz; sequential and simultaneous) resulted in significant modification in the alpha-helix (reduction), beta-turn (reduction), beta-sheet (increase), and random coil (increase) of SCP. The structural alterations shown in the circular dichroism (CD), intrinsic fluorescence spectra, scanning electron microscopy (SEM), surface hydrophobicity, and atomic force microscopy (AFM) of pretreated SCP exhibited the extent of modification caused by cavitation and ultrasonic shear force on enzyme-substrate interaction. Hence, ultrasonic-visco reduction can be applied by industries for enzymatic reaction process enhancement.

Automated summary

The study demonstrated that slit dual-frequency ultrasonic (SDFU) pretreatment significantly enhanced the enzymatic reaction rate of sea cucumber protein (SCP) substrate and reduced the energy cost required for conversion by reducing enthalpy, entropy, and activation energy. Ultrasonic effects also led to significant structural modifications in SCP, impacting enzyme-substrate interactions.