Isolation, Identification, and Characterization of L-asparaginase-Producing Human Commensal Bacterial Strains: A Promising Next-Gen Probiotics.

L-asparaginase is an FDA-approved drug for treating blood cancer, but its inherent antigenicity and L-glutaminase activity are associated with hypersensitivity and organ toxicity. Extracellularly produced glutaminase-free L-asparaginase from human commensal bacteria may be a good alternative to reduce the side effects of therapeutic L-asparaginase. Here, we report the isolation and characterization of fourteen L-asparaginase-producing bacterial strains belonging to the genera Acinetobacter, Escherichia, Klebsiella, and Pseudomonas from human stool and saliva samples. To the best of our knowledge, this is the first report of L-asparaginase-producing human commensal bacterial strains isolated from healthy individuals. L-asparaginase produced by fecal and salivary isolates exhibited significantly higher activity (3.64 to 16.96 U/ml) toward L-asparagine than L-glutamine. Interestingly, L-asparaginase from fecal isolates, Escherichia coli strains 3F1 and 3F2 and salivary isolate Klebsiella pneumoniae 3S3, exhibited no L-glutaminase activity. These isolates were also sensitive to all tested antibiotics. Additionally, these three isolates demonstrated tolerance to pH 3.0 (≥ 88% survival) and 0.3% bile (≥ 95% survival), indicating their potential as probiotics. Among these isolates, L-asparaginase from the highest-producing K. pneumoniae 3S3 strain was found to be a homodimer, with native and subunit molecular weights of 110 kDa and 55 kDa, respectively. The purified enzyme can be further explored for its antitumor and immunomodulatory properties. Overall, future research can be expanded to include the use of a pool of human commensal bacteria as genuine and alternative sources of L-asparaginase for effective cancer treatments and cutting-edge next-generation probiotics.