Bacillus Cucumis: A New Source Of L-Methioninase Enzyme With  Potential For Cancer Therapy

Akhila Warrier; K Hara Gopal; Kadambari Jha Anand; E Legha; N S Soumitri; B S Ashrini; Varalakshmi Kilingar Nadumane

doi:10.48165/

Authors

Akhila Warrier Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)
K Hara Gopal Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)
Kadambari Jha Anand Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)
E Legha Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)
N S Soumitri Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)
B S Ashrini Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)
Varalakshmi Kilingar Nadumane Department of Biotechnology, School of Sciences, Jain (Deemed to be University), JC Road, Bengaluru – 560 027, Karnataka (India)

DOI:

https://doi.org/10.48165/

Keywords:

Bacillus cucumis, cancer, L-methioninase, methionine dependency, MTT assay

Abstract

The existing cancer therapies usually target a single pathway, and the associated side effects and higher costs involved in such mono-target therapies have necessitated to find alternative treatment strategies. Methionine dependency is commonly seen in cancer cells cancerwhereas normal cells in absence of methionine can grow on homocysteine, as they possess intact methionine synthase. So reducing the level of cancer cell/plasma methionine with the help of L methioninase enzyme looks a very promising therapy to manage the cancer. The present research was based on rapid screening of soil samples for L methioninase producing microbes using selective media. The selected microbes were tested for their L-methioninase activity and total protein content. The results indicated a new strain of Bacillus, identified as Bacillus cucumis LAKHS12, exhibiting the highest L-methioninase activity. The cytotoxic activity on in-vitro cancer cell lines by MTT assay revealed that L-methioninase from B. cucumis exhibited highest cytotoxicity. The percentage viability of cervical cancer Hela cells was 35.7%, indicating a significant cytotoxicity of 64.3% for 48 h of treatment with L-methioninase at a lowconcentration. This finding was further supported by the microscopic observations of treated cancer cells. It can be concluded that the isolate has potential for further enzyme purification, characterization, and anticancer applications.

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References

Alshehri, W.A. 2020. Bacterium Hafnia alvei secretes L-methioninase enzyme: Optimization of the enzyme secretion conditions. Saudi Journal of Biological Sciences, 27(5): 1222-1227. Al-Zahrani, S.H.M. and Bukhari, K.A. 2019. Genetics identification and detection of L-methioninase gene in a pseudomonas isolate. Pharmacophore, 10(6): 93-98.

Aneja, K.R. 2003. Experiments in Microbiology and Plant Pathology. New Age International Pvt. Ltd., New Delhi, India.

Arfi, K. 2003. Catabolism of volatile sulfur compounds precursors by Brevibacterium linens and Geotrichum candidum, two microorganisms of the cheese ecosystem. Journal of Biotechnology, 105(3): 245-253.

Bhawana, K. and Priyanka, S. 2018. Microbial production of L-methioninase and its biotechnology application. International Journal of Recent Scientific Research, 9(8C): 28439-28446. Bopaiah, B.B.K., Kumar, D.A.N., Balan, K., Dehingia, L., Reddy, M.K.R.V., Suresh, A.B. and Nadumane, V.K. 2020. Purification, characterization, and antiproliferative activity of L methioninase from a new isolate of Bacillus haynesii JUB2. Journal of Applied Pharmaceutical Sciences, 10(10): 54-61.

Cellarier, E., Durando, X., Vasson, M., Farges, M., Demiden, A., Maurizis, J., Madelmont, J. and Chollet, P. 2003. Methionine dependency and cancer treatment. Cancer Treatment Reviews, 29(6): 489-499.

El-Sayed, A.S. 2010. Microbial L-methioninase: Production, molecular characterization, and therapeutic applications. Applied Microbiology and Biotechnology, 86(2): 445-467.

Akhila Warrier et al.

El-Sayed, A.S. 2009. L-methioninase production by Aspergillus flavipes under solid-state fermentation. Journal of Basic Microbiology, 49(4): 331-341.

Gomez, A. and Gomez, A. 1984. Statistical Procedure for Agricultural Research. (2nd edn.). Kwanchi: John Willy and Sons, New York, USA.

Guo, H., Lishko, V.K., Herrera, H., Groce, A., Kubota, T. and Hoffman, R.M. 1993. Therapeutic tumor-specific cell cycle block induced by methionine starvation in vivo. Cancer Research, 53: 5676-5679.

Hoffman, R.M. 1984. Altered methionine metabolism, DNA methylation and oncogene expression in carcinogenesis. Biochimica et Biophysica Acta (BBA) - Reviews on Cancer, 738(1-2): 49-87. Hoffman, R.M. and Jacobsen, S.J. 1980. Reversible growth arrest in simian virus 40-transformed

human fibroblasts. Proceedings of the National Academy of Sciences, 77(12): 7306-7310. Hori, H., Takabayashi, K., Orvis, L., Carson, D.A. and Nobori, T. 1996. Gene cloning and characterization of Pseudomonas putida L-methionine-α-deamino-γ-mercaptomethane-lyase. Cancer Research, 56(9): 2116-2122.

Ito, S., Nakamura, T. and Eguchi, Y. 1975. Exchange reactions catalysed by methioninase from Pseudomonas putida. The Journal of Biochemistry, 78(5): 1105-1107.

Kahraman, H., Aytan, E. and Kurt, A.G. 2011. Production of methionine γ-lyase in recombinant Citrobacter freundii bearing the hemoglobin gene. BMB Reports, 44(9): 590-594. Kavya, D. and Nadumane, V.K. 2020. Identification of highest L-methioninase enzyme producers among soil microbial isolates, with potential antioxidant and anticancer properties. Journal of Applied Biology and Biotechnology, 8(6): 21-27.

Kavya, D. and Nadumane, V.K. 2023. Enhanced l-methioninase production by Methylobacterium sp. JUBTK33 through RSM and its anticancer potential. Biocatalysis and Agricultural Biotechnology, 47: 102621 [doi.org/10.1016/j.bcab.2023.102621].

Kokkinakis, D.M., Schold, S.C., Hori, H. and Nobori, T. 1997. Effect of long‐term depletion of plasma methionine on the growth and survival of human brain tumor xenografts in athymic mice. Nutrition and Cancer, 29(3): 195-204.

Kreis, W. and Hession, C. 1973. Isolation and purification of L-methionine-α-deamino-γ-mercapto methanelyase (L-methioninase) from Clostridium sporogenes. Cancer Research, 33: 1862-1865. Kumar. S., Stecher, G., Li, M., Knyaz, C. and Tamura, K. 2018. MEGA X: Molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution, 35(6): 1547- 1549.

Laakso, S. and Nurmikko, V. 1976. A spectrophotometric assay for demethiolating activity. Analytical Biochemistry, 72(1-2): 600-605.

Manukhov, I.V., Mamaeva, D.V., Rastorguev, S.M., Faleev, N.G., Morozova, E.A., Demidkina, T.V. and Zavilgelsky, G.B. 2005. A gene encoding L-methionine γ-lyase is present in Enterobacteriaceae family genomes: Identification and characterization of Citrobacter freundii L-methionine γ-lyase. Journal of Bacteriology, 187(11): 3889-3893.

McDonald, D., Price, M.N., Goodrich, J., Nawrocki, E.P., DeSantis, T.Z., Probst, A., Andersen, G.L., Knight, R. and Hugenholtz, P. 2012. An improved green genes taxonomy with explicit ranks for ecological and evolutionary analyses of bacteria and archaea. The ISME Journal, 6(3): 610-618.

Mignard, S. and Flandrois, J.P. 2006. 16S rRNA sequencing in routine bacterial identification: A 30- month experiment. Journal of Microbiological Methods, 67(3): 574-581.

Mosmann, T. 1983. Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. Journal of Immunological Methods, 65(1-2): 55-63. Nejadi, N., Masti, S.M., Tavirani, M.R. and Golmohammadi, T. 2014. Comparison of three routine protein precipitation methods: acetone, TCA/acetone wash and TCA/acetone. Archives of Advances in Biosciences, 5(4): 58-60.

Prihanto, A.A. 2018. IOP Conference Series: Earth and Environmental Science, 137: 012077. [doi:10.1088/1755-1315/137/1/012077].

Anticancer L-methioninase from B. cucumis 399

Sadraeian, M., Rasoul-Amini, S., Mansoorkhani, M.J.K., Mohkam, M., Ghoshoon, M.B. and Ghasemi, Y. 2013. Induction of antitumor immunity against cervical cancer by protein HPV-16 E7 infusion with ricin B chain in tumor-bearing mice. International Journal of Gynecological Cancer, 23(5): 809-814.

Salim, N.A. and Santhiagu, K. Joji. 2019. Process modeling and optimization of high yielding L methioninase from a newly isolated Trichoderma harzianum using response surface methodology and artificial neural network coupled genetic algorithm. Biocatalysis and Agricultural Biotechnology, 17: 299-308.

Selim, M.H, Elshikh, H., El-Hadedy, D., Saad, M.M, Eliwa, E. and Abdelraof, M. 2015a. L methioninase from some Streptomyces isolates I: Isolation, identification of best producers and some properties of the crude enzyme produced. Journal of Genetic Engineering and Biotechnology, 13(2): 129-137.

Selim, M.H., Karm Eldin, E.Z., Saad, M.M., Mostafa, E.S.E., Shetia, Y.H. and Anise, A.A.H. 2015. Purification, characterization of L-methioninase from Candida tropicalis, and its application as an anticancer. Biotechnology Research International, 2015: 1-10. [https://doi.org/10.1155/2015/173140].

Sharma, B., Singh, S. and Kanwar, S.S. 2014. L-methionase: A therapeutic enzyme to treat malignancies. BioMed Research International, 2014: 506287. [doi: 10.1155/2014/506287]. Shirling, E.B. and Gottlieb, D. 1966. Methods for characterization of Streptomyces species, International Journal of Systematic Bacteriology, 16(3): 313-340.

Sundar, W. and Nellaiah, H. 2013. Production of methioninase from Serratia marcescens isolated from soil and its anti-cancer activity against Dalton’s lymphoma ascitic (DLA) and Ehrlich ascitic carcinoma (EAC) in Swiss albino mice. Tropical Journal of Pharmaceutical Research, 12(5): 699-704.

Swisher, E.M., Gonzalez, R.M., Taniguchi, T., Garcia, R.L., Walsh, T. Goff, B.A. and Welcsh, P. 2009. Methylation and protein expression of DNA repair genes: Association with chemotherapy exposure and survival in sporadic ovarian and peritoneal carcinomas. Molecular Cancer, 8(1): 48. [https://doi.org/10.1186/1476-4598-8-48].

Tanaka, H., Esaki, N., Yamamoto, T. and Soda, K. 1976. Purification and properties of methioninase from Pseudomonas ovalis. FEBS Letters, 66(2): 307-311.

Venkatachalam, P. and Nadumane, V.K. 2018. Overexpression of P53 and Bax mediating apoptosis in cancer cell lines induced by a bioactive compound from Bacillus endophyticus JUPR15. Process Biochemistry, 73: 170-179.