Aydın ve Denizli’deki Sıcak Su Kaynaklarından İzole EdilenTermofilik Bakterilerin Lipaz, Amilaz ve Proteaz Aktivitelerinin Araştırılması

Öz

Termofilik bakteriler sıcak çevrelerde gelişebilen mikroorganizmalardır. Bu bakterilerin ürettikleri enzimler bitki ve hayvan kaynaklı enzimlere göre daha stabil olabilmekte ve daha ucuz olarak elde edilmektedirler.Termostabil enzimler ekstrem şartlarda esneklik ve tolerans göstermektedirler. Lipazlar biyoteknoloji ile ilişkili önemli gruplardır ve gıda, mandıra, deterjan ve ilaç endüstrisinde kullanılmaktadırlar. Diğer önemli bir enzim grubu olan amilazlar kağıt, şeker, tekstil, deterjan, fırında pişirme ve demleme endüstrisi gibi çeşitli alanlarda kullanılmaktadır. Proteazlar hidrolitik enzimlerdir ve deterjan üretimi, deri ürünleri, ilaç, et yumuşatıcı olarak, gıda ürünleri ve katı atık arıtma sanayii gibi çeşitli endüstriyel işlemlerde kullanılmaktadır. Bu çalışmada, Aydın ve Denizli’deki bazı termal alanlardan elde edilen 30 suşun 16S rDNA bölgeleri PCR ile çoğaltılmış ve moleküler tanıları yapılmıştır. Otuz izolat arasında suşların %50’si lipaz, %63.3’ü amilaz, %56.6’sı proteaz enzimlerini üretmektedirler. Lipaz-proteaz üreten üç, amilaz-proteaz üreten yedi izolat saptanırken, her üç enzimi de üreten 6 izolat olduğu belirlenmiştir.Sonuç olarak, çalışmamızda elde edilen suşlar ve ürettiklerienzimler endüstriyel biyoteknolojide kullanılma potansiyeline sahiptir.

Anahtar Kelimeler

• Termofilik Bakteriler
• Lipaz
• Amilaz
• Proteaz
• 16S rDNA

Lipase, Amylase and Protease Activities Of Thermophilic Bacteria Isolated From Hot Springs In Aydin And Denizli

Öz

Thermophilic microorganisms are able to grow at high temperatures. Enzymes produced by these kind of bacteria are more stable and cost effective. Thermostable enzymes have flexibility and tolerance in extreme conditions. Lipases dealing with biotechnology are important groups of enzymes. These enzymes are used in food, dairy, detergent and pharmaceutical industry. Amylases that are also important groups are used in various biotechnological fields such as paper, sugar, textile, detergent, baking and infusion industry. Proteases are hydrolytic enzymes and they are used in various industries such as producing detergent, leather products, pharmaceutical, meat tenderizer, food products and solid waste treatment industry. In this study, 16S rDNA regions of thirty bacterial strains isolated from thermal areas in Aydin and Denizli are amplified by PCR and isolates were identified by molecular methods. Among 30 isolates, percentage of strains that produce one enzyme is that; lipase %50, amylase %63.3, protease %56.6. Three of strains were found to be produce both Lipase and protease, while seven strains were amylase and protease producers. Six strains were found to capable of producing all three enzymes. As a result, isolates obtained in our study and their enzymes have an important potential in industrial biotechnology.

Anahtar Kelimeler

• Thermophilic Bacteria
• Lipase
• Amylase
• Protease
• 16S rDNA

Kaynakça

1. ANONİM. Kaplıcaya Sahip Belediyeler Birliği. (2001).
2. Baker, G. C., Gaffar, S., Cowan, D. A., & Suharto, A. R. (2001). Bacterial community analysis of Indonesian hot springs. FEMS Microbiology Letters, 200(1), 103-109.
3. Bakir, Z. B.,& Metin, K. (2016). Purification and characterization of an alkali-thermostable lipase from thermophilic Anoxybacillus flavithermus HBB 134. Journal of microbiology and biotechnology, 26(6), 1087-1097.
4. Başbülbül G., Dabanca M. B., Kaya K. G., Oryaşın. E. ve Bozdoğan. B.(2018) Acıgöl (Denizli)’den İzole Edilen Alkalifilik Bakterilerin Enzimatik Aktivitelerinin Araştırılması. 1st International Health Science And Life Congress (IHSLC 2018).
5. Castro-Ochoa, L. D., Rodríguez-Gómez, C., Valerio-Alfaro, G., & Ros, R. O. (2005). Screening, purification and characterization of the thermoalkalophilic lipase produced by Bacillus thermoleovorans CCR11. Enzyme and Microbial Technology, 37(6), 648-654.
6. Chang, J., Lee, Y. S., Fang, S. J., Park, I. H., & Choi, Y. L. (2013). Recombinant expression and characterization of an organic-solvent-tolerant α-amylase from Exiguobacterium sp. DAU5. Applied biochemistry and biotechnology, 169(6), 1870-1883.
7. Connor, N., Sikorski, J., Rooney, A. P., Kopac, S., Koeppel, A. F., Burger, A., ... & Cohan, F. M. (2010). Ecology of speciation in the genus Bacillus. Applied and environmental microbiology, 76(5), 1349-1358.
8. de Souza, P. M. & e Magalhães, P. de O. Application of microbial α-amylase in industry - a review. Brazilian Journal of Microbiology (2010). doi:10.1590/s1517-83822010000400004

9. Fıtrıanı, S.,& Güven, K. (2018). Isolation, screening, partial purification and characterization of protease from halophilic bacteria isolated from Indonesian fermented food. Anadolu Üniversitesi Bilim ve Teknoloji Dergisi-C Yaşam Bilimleri Ve Biyoteknoloji, 7(2), 130-142.
10. Güven, R. G. (2011). Termofilik bakteriler ve biyoteknolojik açıdan önemli bazı enzimleri. Elektronik Mikrobiyoloji Dergisi TR, 9(1), 1-10.
11. Lessuise, E., Schanck, K., & Colson, C. (1993). Purification and primary characterization of the extracellular lipase of Bacillus subtilis 168, an extreme basic tolerant enzyme. Eur. J. Biochem, 216, 155-160.
12. Mohammad, B. T., Al Daghistani, H. I., Jaouani, A., Abdel-Latif, S., & Kennes, C. (2017). Isolation and characterization of thermophilic bacteria from Jordanian hot springs: Bacillus licheniformis and Thermomonas hydrothermalis isolates as potential producers of thermostable enzymes. International journal of microbiology, 2017.
13. Nawani, N.,& Kaur, J. (2007). Studies on lipolytic isoenzymes from a thermophilic Bacillus sp.: Production, purification and biochemical characterization. Enzyme and Microbial Technology, 40(4), 881-887.
14. Royter, M., Schmidt, M., Elend, C., Höbenreich, H., Schäfer, T., Bornscheuer, U. T., & Antranikian, G. (2009). Thermostable lipases from the extreme thermophilic anaerobic bacteria Thermoanaerobacter thermohydrosulfuricus SOL1 and Caldanaerobacter subterraneus subsp. tengcongensis. Extremophiles, 13(5), 769-783.
15. Ramnath, L., Sithole, B., & Govinden, R. (2017). Identification of lipolytic enzymes isolated from bacteria indigenous to Eucalyptus wood species for application in the pulping industry. Biotechnology Reports, 15, 114-124.
16. Rekik, H., Jaouadi, N. Z., Gargouri, F., Bejar, W., Frikha, F., Jmal, N., ... & Jaouadi, B. (2019). Production, purification and biochemical characterization of a novel detergent-stable serine alkaline protease from Bacillus safensis strain RH12. International journal of biological macromolecules, 121, 1227-1239.
17. Sulong, M. R., Rahman, A., & RN, S. AB, & Basri, MA (2006). Novel organic solvent tolerant lipase from Bacillus sphaericus 205y: extracellular expression of a novel OST-lipase gene. Protein Expression and Purification, 49(2), 190195.