Microstructural, physicochemical properties, and interaction mechanism of hydrogel nanoparticles modified by high catalytic activity transglutaminase crosslinking

Natural protein hydrogels are commonly used in the food industry due to their versatility, making it crucial to develop hydrogels with superior properties. In this study hydrogel nanoparticles were modified with high-performance transglutaminase obtained by cavity engineering strategy. The optimal mutant, R57L/Y198F/F259W, exhibited a 3.3-fold increases (up to 103.26 U mg−1) in catalytic activity and a 3.3 °C increased in TmE value, resulting in a hydrogel with higher hardness, springiness, cohesiveness, and resilience. The hydrophilic capacity of the hydrogel, such as water holding capacity, swelling rate, and freeze-thaw stability (240 h), were also enhanced by 3.00%, 24.56%, and 9.00%, respectively. Fourier transform infrared spectroscopy (FTIR) analysis indicated a 10.76% increase in α-helix and 9.69% increase in β-sheet contents, which led to improved thermostability and textural properties. Scanning electron microscope (SEM) revealed that the structure of modified hydrogel nanoparticles had a denser, mostly polygonal honeycomb structure. Molecular dynamics (MD) simulations indicated that the mutation altered the secondary structure of C64 from a helix to a loop, which improved catalytic activity, resulting in the formation of more cross-linked structures. The W259 residue was found to play a crucial role in stabilizing the complexes. These findings provide valuable insight for the application of protein-based hydrogels in the food industry.