Optimization of L-asparaginase type II produced by Bacillus velezensis SE۱۱۴

Once the acute lymphoblastic leukemia cells that need L-asparagine are exposed to L-ASNase, they die because of the limitations of L-asparagine. The globally rising rate of ALL also requires extraordinary efforts to discover new microorganisms with high L-ASNase production and efficiency. The aim of this study is the high amount of L-ASNase production. After isolation, the L-ASNase production was optimized using the response surface methodology and the central composite design. Then, in-silico studies were predicted for the L-ASNase-producing gene. In this study, Bacillus velezensis was isolated as an L-ASNase producer from slaughterhouse effluent using the M۹ medium. The optimization process further illustrated Tween ۲۰, glucose, temperature, and L-asparagine, which were more significant for L-ASNase production. Based on statistical prediction by response surface methodology, more enzyme activity (۷.۱۱ U/mL) could be realized at ۰.۶% Tween ۲۰, ۱.۷% glucose, ۵۵°C temperature, and ۱.۸% L-asparagine. The in-silico studies also established that the binding site is located at the N-terminal domain and the active site flexible loop. Additionally, it contained Thr۳۶, Ala۴۷, Tyr۵۰, Glu۸۴, Asp۱۱۷, Thr۱۱۶, Met۱۴۲, Lys۱۸۹, Thr۱۹۳, and Thr۱۹۲ as the conserved and functional residues in L-ASNase. It was concluded that B. velezensis SE۱۱۴  produced L-ASNase type II in the present study. The statistical optimization results also showed that Tween ۲۰, glucose, temperature, and L-asparagine were significant variables affecting the L-ASNase production. In addition, temperature and L-asparagine had noteworthy interactions.

Keywords:

L-Asparaginase , Bacillus velezensis , Optimization , Response surface methodology , Central Composite Design