Effect of Media Composition on the Activity of L-Asparaginase Enzyme Produced by Enterobacter Cloacae Complex Sp. V1 Strain

Abstract

Objective: In this study, it was aimed to investigate the effect of medium composition on the ability of producing L-asparaginase enzyme and the activity of L-asparaginase enzyme isolated from soil samples collected from Van Lake Basin. In addition, it was aimed to determine the location of L-asparaginase positive isolates in bacterial taxonomy based on phenotypic and 16S rDNA sequencing analysis. Methods: Luria-Bertani (LB) Agar medium was used to isolate and cultivate bacteria from soil samples. L-asparaginase-producing isolates were screened for M9 minimal salt medium with 0.5% L-asparagine and 0.001% phenol red. The activity of the enzyme produced by positive isolates was determined by spectrophotometric method at 436 nm. The identification of the L-asparaginase producing isolate was performed according to standard microbiological methods and 16S rDNA sequencing analysis. Results: Of the 10 isolates, only one was clearly positive for L-asparaginase in the M9 minimal salt medium containing L-asparagine and phenol red. The L-asparaginase enzyme produced by this isolate was incubated at a temperature of 35 C 16.02 IU/mL at a pH of 8.0 15.25 IU/mL for a 48 hour incubation period 16.02 IU / mL, in the presence of mannitol as a carbon source 15.69 IU/mL, in the presence of yeast extract 14.55 IU/mL and arginine amino acid 17.63 IU/mL as a source of organic nitrogen. Morphological, physiological and biochemical properties of this isolate were determined. In addition, L-asparaginase positive isolate was found to belong to E. cloacae complex strain according to 16S rDNA sequencing analysis and its status in bacterial taxonomy was determined. Conclusion: This research; In our country, it is one of the first studies about screening of L-asparaginase producing bacteria isolated from nature. In the treatment of ALL, L-asparaginase E. cloacae complex sp. V1 strain, positive for the antineoplastic L-asparaginase enzyme is extremely important for our national gene sources. This strain has been found to have an activity that could potentially be used in the industrial production of the enzyme L-asparaginase. © 2020 Refik Saydam National Public Health Agency (RSNPHA.