Enhancing keratinase production of a native Bacillus paralicheniformis FUM-2 through random mutagenesis using a chemical agent and ultraviolet

Document Type : Research Paper

Authors

1 Department of Biology, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran

2 Department of Chemistry, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran

Abstract

Daily accumulation of feather waste due to the development of the poultry industry is a critical issue, considering the environmental and health hazards. Among feather waste disposal methods, biodegradation is a practical, cheap, and environment-friendly method. This study aimed to enhance the keratinolytic activity of FUM-2 isolate, a native keratinolytic bacterium, using mutations and culture improvement. The isolate was identified as Bacillus paralicheniformis based on biochemical, morphological, and molecular analysis. Using the one factor at a time (OFAT) method, the isolate produced more keratinase at 45 °C, pH 11, 2% carbon substrate concentration, 75% aeration rate, and 48 hours incubation. Also, it was able to degrade both α and β keratin. Culture improvement increased enzyme production from 507.6 U/mL to 1706.4 U/mL. In random mutation using ultraviolet radiation, three mutants were isolated, and a 10-minute UV-treated mutant of UVF2-D showed a 36.3% increase in keratinolytic activity compared to its parent. In the next round of mutagenesis by ethidium bromide, seven double mutants were isolated from the UVF2-D mutant, among which the EUF2-D2 double mutant showed a 79% increase in keratinolytic activity. No viable mutants were isolated from the wild strain using ethidium bromide. Our results show that many organisms can potentially produce beneficial products that could be improved using random mutagenesis and optimization methods, which are more useful for strain improvement than complex and costly molecular techniques. 

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References

Abd El-Aziz, N. M., Khalil, B. E., & Ibrahim, H. F. (2023). Enhancement of feather degrading keratinase of Streptomyces swerraensis KN23, applying mutagenesis and statistical optimization to improve keratinase activity. BMC Microbiology, 23(1), 158. https://doi.org/10.1186/s12866-023-02867-0
Abdel-Fattah, A. M., El-Gamal, M. S., Ismail, S. A., Emran, M. A., & Hashem, A. M. (2018). Biodegradation of feather waste by keratinase produced from newly isolated Bacillus licheniformis ALW1. Journal of Genetic Engineering and Biotechnology, 16(2), 311–318. https://doi.org/10.1016/j.jgeb.2018.05.005
Bapiraju, K., Sujatha, P., Ellaiah, P., & Ramana, T. (2004). Mutation induced enhanced biosynthesis of lipase. African Journal of Biotechnology, 3(11), 618–621.
Bhari, R., Kaur, M., & Singh, R. S. (2021). Chicken feather waste hydrolysate as a superior biofertilizer in agroindustry. Current Microbiology, 78(6), 2212–2230. https://doi.org/10.1007/s00284-021-02491-z
Bokveld, A., Nnolim, N. E., Digban, T. O., Okoh, A. I., & Nwodo, U. U. (2021). Chryseobacterium aquifrigidense keratinase liberated essential and nonessential amino acids from chicken feather degradation. Environmental Technology, 1–11. https://doi.org/10.1080/09593330.2021.1969597
Borhani, M. S., Etemadifar, Z., Biria, D., & Jorjani, E. (2019). Molecular detection and optimization production of an alkaline metalloprotease from Exiguobacterium sp. MSB42. Advanced Research in Microbial Metabolites & Technology, 2, 27–39. https://doi.org/10.22104/armmt.2019.3641.1030
Brandelli, A., Daroit, D. J., & Riffel, A. (2010). Biochemical features of microbial keratinases and their production and applications. Applied Microbiology and Biotechnology, 85(6), 1735–1750. https://doi.org/10.1007/s00253-009-2398-5
Chaisemsaeng, P., Sabu, A., Ansanan, S., & Sirisan, S. (2017). Feather degradation and keratinase production by Bacillus sp. and Lactobacillus sp. International Journal of Biotechnology Research, 7, 29–36.
Daroit, D. J., & Brandelli, A. (2014). A current assessment on the production of bacterial keratinases. Critical Reviews in Biotechnology, 34(4), 372–384. https://doi.org/10.3109/10242422.2010.532549
de Paiva, D. P., de Oliveira, S. S. A., Mazotto, A. M., Vermelho, A. B., & de Oliveira, S. S. (2019). Keratinolytic activity of Bacillus subtilis LFB-FIOCRUZ 1266 enhanced by whole-cell mutagenesis. 3 Biotech, 9(2), 1–12. https://doi.org/10.1007/s13205-018-1527-1
Govarthanan, M., Selvankumar, T., Selvam, K., Sudhakar, C., Aroulmoji, V., & Kamala-Kannan, S. (2015). Response surface methodology-based optimization of keratinase production from alkali-treated feather waste and horn waste using Bacillus sp. MG-MASC-BT. Journal of Industrial and Engineering Chemistry, 27, 25–30. https://doi.org/10.1016/j.jiec.2014.12.022
Govarthanan, M., Selvankumar, T., & Arunprakash, S. (2011). Production of keratinolytic enzyme by a newly isolated feather degrading Bacillus sp. from chick feather waste. International Journal of Pharma and Bio Science, 2(3), 259–265.
Gupta, R., Sharma, R., & Beg, Q. K. (2013). Revisiting microbial keratinases: Next generation proteases for sustainable biotechnology. Critical Reviews in Biotechnology, 33(2), 216–228. https://doi.org/10.3109/07388551.2012.685051
Haki, G., & Rakshit, S. (2003). Developments in industrially important thermostable enzymes. Bioresource Technology, 89(1), 17–34. https://doi.org/10.1016/S0960-8524(03)00033-6
Jaouadi, N. Z., Rekik, H., Elhoul, M. B., Rahem, F. Z., Hila, C. G., Aicha, H. S. B., Badis, A., Toumi, A., Bejar, S., & Jaouadi, B. (2015). A novel keratinase from Bacillus tequilensis strain Q7 with promising potential for the leather bating process. International Journal of Biological Macromolecules, 79, 952–964. https://doi.org/10.1016/j.ijbiomac.2015.05.038
Javed, S., Meraj, M., Bukhari, S., Irfan, R., & Mahmood, S. (2013). Hyper-production of alkaline protease by mutagenic treatment of Bacillus subtilis M-9 using agro-industrial wastes in submerged fermentation. Journal of Microbial and Biochemical Technology, 5, 74–80. https://doi.org/10.4172/1948-5948.1000103
Kumar, J., & Mahal, S. (2021). Isolation of Chrysosporium indicum from poultry soil for keratinase enzyme, its purification and partial characterization. Journal of Applied and Natural Science, 13(2), 744–751. https://doi.org/10.31018/jans.v13i2.2609
Lateef, A., Adelere, I. A., & Gueguim-Kana, E. B. (2015). The biology and potential biotechnological applications of Bacillus safensis. Biologia, 70, 411–419. https://doi.org/10.1515/biolog-2015-0062
Lateef, A., Adelere, I. A., & Gueguim-Kana, E. B. (2015). Bacillus safensis LAU 13: A new source of keratinase and its multi-functional biocatalytic applications. Biotechnology and Biotechnology Equipment, 29(1), 54–63. https://doi.org/10.1080/13102818.2014.986360
Mariyam, I. (2011). Multistep mutagenesis for the over-expression of cellulases in Humicola insolens. Pakistan Journal of Botany, 43(1), 669–677.
McKittrick, J., Chen, P. Y., Bodde, S. G., Yang, W., Novitskaya, E. E., & Meyers, M. A. (2012). The structure, functions, and mechanical properties of keratin. JOM, 64(4), 449–468. https://doi.org/10.1007/s11837-012-0302-8
Mehta, R. S., Jholapara, R. J., & Sawant, C. S. (2014). Isolation of a novel feather-degrading bacterium and optimization of its cultural conditions for enzyme production. International Journal of Pharmacy and Pharmaceutical Sciences, 6(1), 194–201.
Mini, K., Paul, M., & Mathew, J. (2015). Optimization of conditions for production of Keratinase by Aspergillus flavus by submerged fermentation. International Journal of Pharmacy and Bio Science, 6(4), B377–B385.
Moridshahi, R., Bahreini, M., Sharifmoghaddam, M. R., & Asoodeh, A. (2020). Biochemical characterization of an alkaline surfactant-stable keratinase from a new keratinase producer, Bacillus zhangzhouensis. Extremophiles, 24, 693–704. https://doi.org/10.1007/s00792-020-01187-9
Moridshahi, R., Sharifmoghadam, M. R., & Bahreini, M. (2021). Biodegradation of keratinous wastes by alkaline keratinase produced from Bacillus tequilensis BK206 and optimizing the cultural conditions for increased keratinolytic activity. Journal of Applied Biological Science, 15(2), 185–195.
Nnolim, N. E., Udenigwe, C. C., Okoh, A. I., & Nwodo, U. U. (2020). Microbial keratinase: Next generation green catalyst and prospective applications. Frontiers in Microbiology, 11, 580164. https://doi.org/10.3389/fmicb.2020.580164
Pahua-Ramos, M., Hernández-Melchor, D., Camacho-Pérez, B., & Quezada-Cruz, M. (2017). Degradation of chicken feathers. BioTechnology: An Indian Journal, 13(6), 1–24.
Pandey, S. C., Pande, V., Sati, D., Gangola, S., Kumar, S., Pandey, A., & Samant, M. (2019). Microbial keratinase: A tool for bioremediation of feather waste. In P. Bhatt (Ed.), Smart Bioremediation Technologies (pp. 217–253). Academic Press. https://doi.org/10.1016/B978-0-12-818307-6.00013-5
Parinayawanich, S., Sittipol, D., Rodpan, S., Pattanapanyasat, K., & Jongruja, N. (2021). Application of recombinant hyperthermostable keratinase for degradation of chicken feather waste. Biocatalysis and Agricultural Biotechnology, 36, 102146. https://doi.org/10.1016/j.bcab.2021.102146
Parekh, S., Vinci, V., & Strobel, R. (2000). Improvement of microbial strains and fermentation processes. Applied Microbiology and Biotechnology, 54(3), 287–301. https://doi.org/10.1007/s002530000403
Purchase, D. (2016). Microbial keratinases: Characteristics, biotechnological applications and potential. In The Handbook of Microbial Bioresources (pp. 634–674). CABI. http://www.cabi.org/bookshop/book/9781780645216
Rodrigers da-Silva, R. (2018). Keratinases as an alternative method designed to solve keratin disposal on the environment: Its relevance on agricultural and environmental chemistry. Journal of Agricultural and Food Chemistry, 66(28), 7219–7221. https://doi.org/10.1021/acs.jafc.8b03152
Salwan, R., & Sharma, V. (2019). Trends in extracellular serine proteases of bacteria as detergent bioadditive: Alternate and environmental friendly tool for detergent industry. Archives of Microbiology, 201, 863–877. https://doi.org/10.1007/s00203-019-01662-8
Sanghvi, G., Patel, H., Vaishnav, D., Oza, T., Dave, G., Kunjadia, P., & Sheth, N. (2016). A novel alkaline keratinase from Bacillus subtilis DP1 with potential utility in cosmetic formulation. International Journal of Biological Macromolecules, 87, 256–262. https://doi.org/10.1016/j.ijbiomac.2016.02.067
Singh, I., & Kushwaha, R. (2015). Keratinases and microbial degradation of keratin. Advances in Applied Science Research, 6(2), 74–82.
Soliman, E. A., El-Hamshary, O., & Batayyib, R. S. (2016). Enhancement of protease production of some Bacillus spp. isolated from various regions in Jeddah City. International Journal of Current Microbiology and Applied Sciences, 5(5), 619–635. http://dx.doi.org/10.20546/ijcmas.2016.505.063
Suribabu, K., Govardha, T., & Hemalatha, K. (2014). Strain improvement of Brevibacillus borostelensis R1 for optimization of α-amylase production by mutagens. Journal of Microbial and Biochemical Technology, 6(3), 123–127. https://doi.org/10.4172/1948-5948.1000132
Tamreihao, K., Mukherjee, S., Khunjamayum, R., Devi, L. J., Asem, R. S., & Ningthoujam, D. S. (2019). Feather degradation by keratinolytic bacteria and biofertilizing potential for sustainable agricultural production. Journal of Basic Microbiology, 59, 4–13. https://doi.org/10.1002/jobm.201800434
Tuly, J. A., Ma, H., Zabed, H. M., Dong, Y., Janet, Q., Golly, M. K., Feng, L., Li, T., & Chen, G. (2021). Harnessing the keratinolytic activity of Bacillus licheniformis through random mutagenesis using ultraviolet and laser irradiations. Applied Biochemistry and Biotechnology, 120. https://doi.org/10.1007/s12010-021-03697-4
Vidmar, B., & Vodovnik, M. (2018). Microbial keratinases: Enzymes with promising biotechnological applications. Food Technology and Biotechnology, 56(3), 312–328. https://doi.org/10.17113/ftb.56.03.18.5658
Zhang, X. (2011). Notice of Retraction: Research on improving keratinase vitality of Bacillus subtilis based on compound mutation. In Proceedings of the 2011 5th International Conference on Bioinformatics and Biomedical.
Microbiology, Metabolites and Biotechnology
Volume 6, Issue 2
November 2023
Pages 63-72

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