Production and optimization of D-amino acid oxidase which is involved in the biosynthesis of β-lactam antibiotics

A total of sixty seven fungal isolates representing 40 fungal species related to 14 genera were screened for their abilities to produce endocellular D-amino acid oxidase enzyme. The most active fungal isolates were Fusarium heterosporum and Nectria haematococca, producing 210.41 and 207.94 units/ml, respectively. Maximum activity of D-amino acid oxidase produced by F. heterosporum and N. haematococca was obtained after 7 days of incubation at 30 degrees C with pH 7 culture medium containing glucose and ammonium sulphate as carbon and nitrogen sources, respectively. Inoculation of cultures by 3 discs of fungi and incubation of cultures at 160 rpm shaking condition improved the enzyme production. Among seven amino acids tested, D-alanine was the best inducer for D-amino acid oxidase production by F. heterosporum; however L-asparagine was the best by N. haematococca. High-performance liquid chromatography (HPLC) analysis indicated that the purified D-amino acid oxidase produced by both fungi was active for the conversion of cephalosporin C to glutaryl-7-aminocephalosporanic acid.