Potential Application of N-Carbamoyl-Alanine Amidohydrolase from Agrobacterium tumefaciens C58 for beta-amino acid Production,

An N-carbamoyl-β-alanine amidohydrolase of industrial interest from Agrobacterium tumefaciens C58 (βcarAt) has been characterized. βcarAt is most active at 30°C and pH 8.0 with N-carbamoyl-β-alanine as a substrate. The purified enzyme is completely inactivated by the metal-chelating agent 8-hydroxyquinoline-5-sulfonic acid (HQSA), and activity is restored by the addition of divalent metal ions, such as Mn2+, Ni2+, and Co2+. The native enzyme is a homodimer with a molecular mass of 90 kDa from pH 5.5 to 9.0. The enzyme has a broad substrate spectrum and hydrolyzes nonsubstituted N-carbamoyl-α-, -β-, -γ-, and -δ-amino acids, with the greatest catalytic efficiency for N-carbamoyl-β-alanine. βcarAt also recognizes substrate analogues substituted with sulfonic and phosphonic acid groups to produce the β-amino acids taurine and ciliatine, respectively. βcarAt is able to produce monosubstituted β2- and β3-amino acids, showing better catalytic efficiency (kcat/Km) for the production of the former. For both types of monosubstituted substrates, the enzyme hydrolyzes N-carbamoyl-β-amino acids with a short aliphatic side chain better than those with aromatic rings. These properties make βcarAt an outstanding candidate for application in the biotechnology industry.