In silico and biochemical analysis on a newly isolated Trichoderma asperellum L-asparaginase

Microbial L-asparaginase enzyme has been considered as a good source of potential anticancer drugs. The enzyme purified from different microbes has been shown different biochemical and kinetics properties. Thus, our study aims to investigate L-asparaginase from Trichoderma asperellum (MN960163), including in silico characterization of the L-asparaginase encoding genes as well as the biochemical characters of the purified enzyme. In silico, analysis of the T. asp genome database revealed the presence of six protein encoding genes with N-terminal asparaginase domain that has asparaginase production potential as indicated from their physico-chemical properties, verification of three of them was confirmed. T. asperellum L- asparaginase with a specific activity of 200 U/mg and 61.7 fold purification was achieved by ammonium sulphate precipitation (60%) followed by DEAE-sepharose ion exchange and Sephacryl S-200 gel filtration chromatography. The purified enzyme has a molecular mass of 65 kDa and exhibited its maximum activity at pH 8 and 50 degrees C. The purified enzyme showed high thermal stability up to 50 degrees C and a high affinity to L- asparagine with a Km of 0.35 mM. The activity was significantly enhanced by Cu, K and Ca ions while Fe and EDTA was completely inhibited L-asparginase activity. In conclusion, the current data highlight new possible genes that could be employed as a promising source in asparaginase production for medical purposes. Additionally, the feature of the purified asparaginase enzyme could help to elucidate the unique properties of T. asp asparaginase.