High Salt-Resistant\nUrethanase Degrades Ethyl Carbamate\nin Soy Sauce

Urethanase\nis a promising biocatalyst for degrading carcinogen\nethyl carbamate (EC) in fermented foods. However, their vulnerability\nto high ethanol and/or salt and acidic conditions severely limits\ntheir applications. In this study, a novel urethanase from Alicyclobacillus pomorum (<i>Ap</i>UH)\nwas successfully discovered using a database search. <i>Ap</i>UH shares 49.4% sequence identity with the reported amino acid sequences.\nIt belongs to the Amidase Signature family and has a conserved “K–S–S”\ncatalytic triad and the characteristic “GGSS” motif.\nThe purified enzyme overexpressed in Escherichia coli exhibits a high EC affinity (<i>K</i><sub>m</sub>, 0.306\nmM) and broad pH tolerance (pH 4.0–9.0), with an optimum pH\n7.0. Enzyme activity remained at 58% in 12% (w/v) NaCl, and 80% in\n10% (v/v) ethanol or after 1 h treatment with the same ethanol solution\nat 37 °C. <i>Ap</i>UH has no hydrolytic activity toward\nurea. Under 30 °C, the purified enzyme (200 U/L) degraded about\n15.4 and 43.1% of the EC in soy sauce samples (pH 5.0, 6.0), respectively,\nin 5 h. Furthermore, the enzyme also showed high activity toward the\nclass 2A carcinogen acrylamide in foods. These attractive properties\nindicate their potential applications in the food industry.