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l-asparaginases play a crucial role in the treatment of acute lymphoblastic leukemia (ALL), a type of cancer that mostly affects children and teenagers. However, it is common for these molecules to cause adverse reactions during treatment. These downsides ignite the search for novel asparaginases to mitigate these problems. Thus, this work aimed to produce and characterize a recombinant asparaginase from Phaseolus vulgaris (Asp-P). In this study, Asp-P was expressed in Escherichia coli with high yields and optimum activity at 40 °C, pH 9.0. The enzyme K m and V max values were 7.05 mM and 1027 U/mg, respectively. Asp-P is specific for l-asparagine, showing no activity against l-glutamine and other amino acids. The enzyme showed a higher cytotoxic effect against Raji than K562 cell lines, but only at high concentrations. In silico analysis indicated that Asp-P has lower immunogenicity than a commercial enzyme. Asp-P induced biofilm formation by Candida sp. due to sublethal dose, showing an underexplored potential of asparaginases. The absence of glutaminase activity, lower immunogenicity and optimal activity similar to physiological temperature conditions are characteristics that indicate Asp-P as a potential new commercial enzyme in the treatment of ALL and its underexplored application in the treatment of other diseases., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Bruno Anderson Matias da Rocha reports financial support was provided by National Council for Scientific and Technological Development. Bruno Anderson Matias da Rocha reports financial support was provided by Coordination of Higher Education Personnel Improvement. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objective: L-Asparaginase (ASNase) is an enzyme used in the treatment of acute lymphoblastic leukemia (ALL). As the therapeutic ASNases has bacterial origin, severe side effects are associated with its use, among them hypersensitivity and inactivation of the enzyme. In this context, the objective of this work was to produce a recombinant ASNase of bacterial origin in human cells in order to determine the presence and consequences of potential post-translational modifications on the enzyme., Results: Recombinant ASNase was expressed in human cells with a molecular weight of 60 kDa, larger than in Escherichia coli, which is 35 kDa. N-glycosylation analysis demonstrated that the increased molecular weight resulted from the addition of glycans to the protein by mammalian cells. The glycosylated ASNase presented in vitro activity at physiological pH and temperature. Given that glycosylation can act to reduce antigenicity by masking protein epitopes, our data may contribute to the development of an alternative ASNase in the treatment of ALL in patients who demonstrate side effects to currently marketed enzymes.