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Microbial transglutaminase (MTG) is an enzyme widely used in the food industry. Mutiple-site mutagenesis of Streptomyces mobaraensis transglutaminase was performed in Escherichia coli. According to enzymatic assay and thermostability study, among three penta-site MTG mutants (DM01-03), DM01 exhibited the highest enzymatic activity of 55.7 ± 1.4 U/mg and longest half-life at 50 °C (418.2 min) and 60 °C (24.8 min).

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The objective of present work was to construct antibacterial [2-(methacryloyloxy) ethyl] trimethylammonium chloride functionalized reduced graphene oxide/poly(ethylene-co-vinyl alcohol) (MTAC-rGO/EVOH) multilayer barrier films by using layer-by-layer assembly under a parallel electric field. Besides barrier and mechanical properties, the antibacterial activities of the film and cytotoxicity of MTAC-rGO nanosheets were extensively investigated. The functionalization of rGO was achieved by grafting MTAC onto a graphene framework through C (sp)-C bonds. The assembly of MTAC-rGO on the EVOH matrix not only significantly improved film mechanical strength, but also endowed the targeting film with outstanding moisture barrier even under a relative humidity of 99% (e.g., 0.019 g m s atm for (MTAC-rGO/EVOH)) besides good oxygen barrier (e.g., 0.07 cm m d atm for (MTAC-rGO/EVOH)). Among the testing films, MTAC-rGO/EVOH film had the best antibacterial activity, and the activity against S. aureus was better than E. coli. Meanwhile, the cytotoxicity of MTAC-rGO nanosheets was very low. Results suggest that MTAC-rGO/EVOH film may have great potential in food active packaging.