مقايسه و بررسي بيوانفورماتيكي آنزيم اسپاراژيناز بين سويه هاي مختلف پروبيوتيك و ارزيابي فعاليت آنزيم آسپاراژيناز در برخي سويه¬هاي پروبيوتيك

Bioinformatics analysis an‎d comparison of asparaginase enzyme between different strains of probiotics an‎d measurement of asparaginase activity in some probiotics

زيست فناوري

L-asparaginase is an enzyme that is used in various industries such as food, medical an‎d medicine due to its hydrolytic activity. Many organisms such as humans, animals, plants an‎d microorganisms have the ability to produce this enzyme. Extracting this enzyme from microbial sources is easier an‎d more economical than other sources. Probiotics are among the most useful an‎d safe microorganism sources of this enzyme. Probiotic sources prepared for this research were first cultivated in solid modified M9 medium. The strains that have the ability to produce this enzyme change the color of the culture medium to purple due to the production of ammonium, which is a base compound. These strains were cultured in liquid modified M9 medium in order to quantitatively measure asparaginase enzyme activity. Their enzyme activity was calculated every 12 hours using Nesslerʹs reagent. The optimal conditions for the production of asparaginase enzyme with the variables of pH, temperature, carbon source an‎d nitrogen source were investigated for some strains. Then, the lethal effect of asparazinase enzyme of some strains, which was partially purified by precipitation with acetone an‎d using a dialysis bag, was observed on MCF-7 cancer cells. Finally, bioinformatics studies such as investigating protein secondary structure, physicochemical properties, motifs an‎d molecular docking were performed on protein sequences of asparaginase enzyme of probiotic strains. According to the results of this research, color change was observed in the culture medium of six strains of Weizmannia coagulans, Bacillus pumilus KX270718, Bacillus pumilus KX270719, Bacillus cereus, Bacillus subtilis an‎d Bacillus mojavensis. The highest an‎d lowest levels of asparaginase enzyme activity were among the strains of Bacillus pumilus KX270719, Bacillus subtilis, Weizmannia coagulans, Bacillus mojavensis, Bacillus pumilus KX270718 an‎d Bacillus cereus. In order to better investigate the release time of the enzyme, the graph of the asparaginase enzyme activity of the strains was matched with the growth curve of each strain. Asparaginase enzyme of Bacillus pumilus KX270719, Bacillus cereus an‎d Bacillus subtilis strains were released in the logarithmic phase, Weizmannia coagulans an‎d Bacillus mojavensis strains in the stationary phase an‎d Bacillus pumilus KX270718 strain in the death phase. Optimum conditions for the production of asparaginase enzyme with variables of pH, temperature, carbon source an‎d nitrogen source in two strains of Bacillus pumilus KX270719 an‎d Weizmannia coagulans were pH 8.4, temperature 42 an‎d 37 °C, carbon source glucose an‎d nitrogen source yeast extract an‎d peptone respectively. The results of the lethal effect of asparaginase enzyme of Bacillus pumilus KX270719 an‎d Weizmannia coagulans strains showed that the lethal effect of asparaginase enzyme of Bacillus pumilus strain KX270719 is more than the lethal effect of asparaginase enzyme of Weizmannia coagulans strain an‎d in both strains, the lethal effect increases with increasing enzyme concentration.

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