Purification of L-Amino acid oxidase obtained from Aspergillus terreus MZ769058 and its biochemical characterization

Abstract

Abstract L-Amino acid oxidase has significant values in different biotechnology sectors. In this study, Aspergillus terreus MZ769058 had been reported as new fungal isolate for production of this enzyme. It was partially purified using ammonium sulfate precipitation methodology with87.9 U/mg protein maximal specific activity and1.69 folds purification fold for 30–60% saturation. Ion-exchange chromatography was further applied for achieving high value of purification fold (2.55) and high specific activity as 132.5 U/mg.This purified enzyme showed homodimer nature with molecular weight of subunit as 90 kDa and 180 kDa by SDS electrophoresis and NATIVE PAGE respectively. The activity of enzyme was found as maximum 193.5 U/l at optimum value of pH 6.0.The enzyme was active throughout a wide range of temperatures and showed maximum activity (227.08U/l) at optimum temperature 30°C. The value of Michaelis parameters of Km and Vmaxwas estimated as 26 mM and 250 µmole/min/mg proteinsrespectively. The catalytic efficiency of this enzyme (Kcat) value was determined as 2.5 µmole/min/mg. Metal ions such as FeSO4 (85.4 %), Na2MO4 (81.2 %), and CuSO4 had showed negative effect on the activity of LAAO enzyme. Metal ions like MgSO4, H3BO3, and ZnSO4had showed very little effect on activityof L-Amino acid oxidase. The activity of LAAO enzyme was strongly inhibited at a concentration of 10 mM CaCl2.This enzyme was strongly inhibited with α-napthol (34.4%), EDTA (34.2%), Glycine (39%) sodium azide (41.4%), and riboflavin (85.3%). Fourier transform infrared spectroscopyhad confirmed the presence of the amine and aldehyde groups with C-H stretch, C=O stretch, C-O stretch at peak of 2927.95, 1745.25, and 1078.64 cm-1. This enzyme could be effectively used for effective therapeutic agent in pharmaceutical sector.