Optimization of L-asparaginase production using native soil-isolated Bacillus sp. and evaluation of its anticancer activity
Subject Areas : Microbial Biotechnology
Forough Rahnamay Roodposhti
1
(Department of Biology, Faculty of Basic Sciences, Rasht Branch, Islamic Azad University, Rasht, Iran)
Leila Asadpour
2
(Department of Biology, Faculty of Basic Sciences, Rasht Branch, Islamic Azad University, Rasht, Iran)
Mahdi Shahriarinour
3
(Department of Biology, Faculty of Basic Sciences, Rasht Branch, Islamic Azad University, Rasht, Iran)
Behnam Rasti
4
(Department of Microbiology, Faculty of Basic Sciences, Lahijan Branch, Islamic Azad University, Lahijan, Guilan, Iran)
Sajjad Gharaghani
5
(Laboratory of Bioinformatics and Drug Design (LBD), Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran)
Keywords: Fatty Acids, GC-MS, Candida glabrata, Yeast, microbial species,
Abstract :
Background and Objectives: Bacteria are one of the most important sources of L-asparaginase (ASNase) production which is used as an anticancer agent in the treatment of acute lymphoblastic leukemia worldwide. This study aimed to optimize the ASNase production by bacteria isolated from the soil in northern Iran, and to determine its anti-cancer activity. Materials and Methods: ASNase production by bacterial strains isolated from forest soil samples in Guilan Province, northern Iran was investigated. The optimized condition of enzyme production, kinetics, effect of activators and inhibitors and anticancer activity of the partially purified L-asparaginase against MCF-7 cell lines were studied. Results: A promising ASNase producing isolate, was identified by 16S rRNA gene sequencing as Bacillus sp. The glutaminase activity of the enzyme was found to be 5.9 times lower than its asparaginase activity and the enzyme showed affinity for L-asparagine with a Km value and Vmax of 0.055M and 35.71 µM/mL/min, respectively. The current ASNase enzyme was stable from pH 6.5 to 8.5 and stable up to 55°C. ASNase activity was not significantly affected by the presence of two metal ions Na+, K+; Mg2+ showed enhancement in enzyme activity, while Ca2+ decreased it. Anticancer activity of the purified L-asparaginase was detected against MCF-7 cell lines with IC50 of 21µg/ml. Conclusion: The soil isolate Bacillus sp. was identified as a candidate for L-asparaginase production. The future prospect of this enzyme recommends its utility in pharmaceutical and food industry.
References
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28.Hassan SW, Farag AM, Beltagy EA. Purification, characterization and anticancer activity of L-asparaginase produced by marine Aspergillus terreus. J. Pure Appl. Microbiol. 2018;12(4):1845-54.
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_||_References
2.Sindhwad P, Desai K. media optimization, Isolation and purification of L-asparaginase from marine isolate. Asian Pac. J. Health Sci. 2015; 293:72-82.
9.Gomori G. Preparation of buffers for use in enzyme studies. In S. P. Colowick & N. O. Kaplan (Eds.), Methods in Enzymology; 1995: 138–146. New York: Academic Press.
10.El-Naggar NE, Deraz SF, El-Ewasy SM, Suddek GM. Purification, characterization and immunogenicity assessment of glutaminase free L-asparaginase from Streptomyces brollosae NEAE-115. BMC Pharmacology and Toxicology. 2018;19(1):51.
11.Kumari PV, Sankar GG, Prabhakar T, Lakshmi SS. Purification and characterization of L-asparaginase from Streptomyces griseoluteus WS3/1. Int J Pharm Sci Rev Res. 2013;23(2):198-202.
12.Monica T, Lincoln L, Niyonzima FN, Sunil SM. Isolation, purification and characterization of fungal extracellular L-asparaginase from Mucor Hiemalis. J Biocatal Biotransform 2013; 2:1-9.
13.Warangkar SC, Khobragade CN. Purification, characterization, and effect of thiol compounds on activity of the Erwinia carotovora L-asparaginase. Enzyme Res 2010; 1:1-10.
14.Alrumman SA, Mostafa YS, Al-Izran KA, Alfaifi MY, Taha TH, Elbehairi SE. Production and anticancer activity of an L-asparaginase from Bacillus licheniformis isolated from the Red Sea, Saudi Arabia. Scientific reports. 2019;9(1):1-4.
15.Mahajan RV, Kumar V, Rajendran V, Saran S, Ghosh PC, Saxena RK. Purification and characterization of a novel and robust L-asparaginase having low-glutaminase activity from Bacillus licheniformis: in vitro evaluation of anti-cancerous properties. PLoS One. 2014 6;9(6):e99037.
16.Verma N, Kumar K, Kaur G, Anand S. L-asparaginase: a promising chemotherapeutic agent. Critical reviews in biotechnology. 2007;27(1):45-62.
17.Kumar NM, Ramasamy R, Manonmani HK. Production and optimization of l-asparaginase from Cladosporium sp. using agricultural residues in solid state fermentation. Industrial Crops and Products. 2013; 43:150-8.
18.Thenmozhi C, Sankar R, Karuppiah V, Sampathkumar P. L-asparaginase production by mangrove derived Bacillus cereus MAB5: optimization by response surface methodology. Asian Pacific journal of tropical medicine. 2011;4(6):486-91.
20.Mostafa Y, Alrumman S, Alamri S, Hashem M, Al-izran K, Alfaifi M, Elbehairi SE, Taha T. Enhanced production of glutaminase-free L-asparaginase by marine Bacillus velezensis and cytotoxic activity against breast cancer cell lines. Electronic Journal of Biotechnology. 2019 Nov 1;42:6-15.
22.Rahimzadeh M, Poodat M, Javadpour S, Qeshmi FI, Shamsipour F. Purification, characterization and comparison between two new L-asparaginases from Bacillus PG03 and Bacillus PG04. The open biochemistry journal. 2016; 10:35-45.
27.Safary A, Moniri R, Hamzeh-Mivehroud M, Dastmalchi S. Highly efficient novel recombinant L-asparaginase with no glutaminase activity from a new halo-thermotolerant Bacillus strain. BioImpacts: BI. 2019;9(1):15-23.
28.Hassan SW, Farag AM, Beltagy EA. Purification, characterization and anticancer activity of L-asparaginase produced by marine Aspergillus terreus. J. Pure Appl. Microbiol. 2018;12(4):1845-54.
29.Moharib SA. Anticancer activity of L-Asparaginase produced from Vigna uUnguiculata. World Sci. Res. 2018; 5: 1-12.
30.Shafei MS, El-Refai HA, Mostafa H, El-Refai AM, El-Beih FM, Easa SM, Gomaa SK. Purification, characterization and kinetic properties of Penicillium cyclopium L-asparaginase: Impact of lasparaginase on acrylamide content in potato products and its cytotoxic activity. Cur. Tre. Biotech. Pharm. 2015 Apr 1;9:132-40.