SCREENING AND PRODUCTION OF MICROBIAL L-ARGINASE ENZYME AS ANTICANCER AGENT FROM DIFFERENT SOIL ENVIRONMENTS IN EGYPT

Background: Hydrolysis of L-arginine to L-ornithine and urea is catalyzed by the crucial L-arginase enzyme. L-arginase nowadays is necessary for medicine due to its significance as an anticancer agent against auxotrophic cancers to arginine. It exists and is widely distributed in plants, fungi, chordates, and non chordates, however, there are few available reports for L-arginase from bacteria. Auxotrophic cancer cells for L-arginine can not synthesize their L-arginine as a sole metabolic source of carbon and nitrogen, while normal cells can synthesize their L-arginine for normal growth. Deprivation of auxotrophic cancer cells from L-arginine causes their killing. The aim of the study: Our study was aimed at isolation, screening, and molecular detection of L-arginase expressing bacteria from different soil environments in Egypt and characterization of the enzyme of activity as an anticancer mediator against auxotrophic cancer cells for arginine.
Materials and methods: 50Grassland soil samples were collected from different locations in Egypt. The activity of L-arginase was confirmed by rapid plate assay screening using mineral arginine agar (MAA) media and estimating the optical density of Urea level spectrophotometrically. DNA Northen blotting hybridization technique, Gram stain, and biochemical reactions were used to identify the predominant soil bacterial isolates producing L-arginase. Tissue culturing techniques were applied for screening of anticancer activity of bacterial L-arginase against auxotrophic cancers for arginine.
Results: Only the bacterial isolates which were able to utilize L-arginine as the sole metabolic source for carbon and nitrogen were grown on MAA media. Molecular detection and the biochemical reactions revealed that Bacillus subtilis, strain W23 (74%), and Bacillus anthracis, strain 34F2 (26%) were the predominant bacterial isolates producing L-arginase as an anticancer mediator for auxotrophic cancers for arginine. Bacterial L-arginase showed excellent anticancer activity against auxotrophic cancers for L-arginine in presence of Co, Ni, and Mn metal ions as co-factors. The molecular weight of bacterial L-arginase was found to be 39KDa as determined by the mass spectrometer.IC50 of bacterial L-arginase was 2.31 U/ml against hepatic cancer cell line,2.19 U/ml against melanoma cancer cell line and 4.51 U/ml against colorectal cancer cell line,4.64 U/ml against lung adenocarcinoma cell line (Calu-3) and 4.28 U/ml against cardiac cancer line(HL1). The Km and Vmax values of L-arginase were 8.63 mmol/l and 7.41 μmol /min. Conclusion: Our study was a promising approach study due to discovering a new bacterial L-arginase enzyme as an anticancer agent from different soil environments in Egypt.