Green bioprocessing of L-Asparaginase: optimization, purification, and characterization using novel Fusarium solani...

L-Asparaginase (L-ASNase) (EC 3.5.1.1) produced by fungi has tremendous potential in the therapeutic and food processing sectors. In the present study, a newly isolated fungal strain, PVS1, identified as Fusarium solani (GenBank accession no. PQ721280; Indian Type Culture Collection no. ITCC 9504), produced a high amount of extracellular L-ASNase, 44.6 U/mL, on the 4th day in the modified Czapek Dox medium under submerged conditions. The production of L-ASNase was optimized using the One-Factor-at-A-Time (OFAT) approach, which demonstrated that the most suitable carbon and nitrogen sources were glycerol (2.0 g/L) and peptone (10.0 g/L), respectively, amended with L-asparagine (5.0 g/L). Additionally, F. solani PVS1 demonstrated high production of L-ASNase at a pH of 6.0 ± 0.2, incubation temperature of 37°C, and under shaking conditions, resulting in a maximum activity of 58.8 U/mL on the 3rd day. Partial purification of the enzyme was achieved through acetone precipitation and diethylaminoethyl (DEAE)-Sephadex ion exchange chromatography, resulting in a 3.53-fold purification with a recovery rate of 0.28%. The partially purified L-ASNase exhibited a molecular weight of approximately 70 kDa, with activity over an extended temperature range from 20 to 60 °C, and showed the maximum residual activity of 50.3% at pH 5.0 ± 0.2. Furthermore, L-ASNase exhibited stability with most of the tested metal ions. The K_m and V_max of the L-ASNase were 7.14 mM and 102.14 U/mL, respectively. The partially purified L-ASNase exhibited a high DPPH scavenging activity (IC₅₀ 0.51 ± 0.02 mg/mL). In conclusion, the results demonstrate that the F. solani PVS1 strain is a prospective candidate for producing L-ASNase and could be applied for subsequent applications in the pharmaceutical and food processing industries.