Glutaminase free L-asparaginase from seaweed associated Bacillus licheniformis with promising therapeutic potential

The present study highlights the detailed investigation of the physico-chemical characteristics of the anti-cancer enzyme L-aspraginase (LA) from the red seaweed Gracilaria dura-associated Bacillus licheniformis strain. We performed the enzyme purification, cytotoxicity assays, toxicity analysis on Caenorhabditis elegans, and kinetics characterisation. We standardized a two-step enzyme purification strategy, involving ammonium sulfate precipitation followed by Sephadex column chromatography in this study to achieve high enzyme purity. The purified LA exhibited high specificity for L-asparagine with a low Km (0.014 ± 0.0006 mM), absence of L-glutaminase activity; and a high level of enzyme activity (229 IU ml−1). This LA variant demonstrated cytotoxicity against HEK 293 cells and it did not affect the normal growth and life span of C. elegans. There fore this study underpins the possibility of utilising this LA variant for clinical applications. The enzyme is functional at a wide pH and temperature range, with optimal conditions aligning with the physiological levels of human blood (pH 7.37 °C). Moreover, the enzyme is secreted extracellularly leading to a simple downstream processing. We assume that the potential immunogenecity of this enzyme in humans to be minimal or absent, however it should be substantiated through assessments on higher organisms. Our findings not only provides useful insights into the potential of marine LA in human therapeutics but also shed light into the possibility of obtaining high value products from seaweed associated bacteria. This study enhances the scope of carrying out research on the seaweed ecology and interactions in the marine environment.