Immobilize Pleurotus ostreatus ve Lentinus tigrinus Misellerinin Birlikte Kültürü ile Lakkaz ve Mangan Peroksidaz Enzim Aktivitelerinin Arttırılması

Öz

Bu çalışmada lignin modifiye edici enzimlerin fazla miktarda üretilmesi için immobilize fungus misellerinin birlikte kültür edilmesi ilk kez olarak rapor edilmiştir. Pleurotus ostreatus OBCC 6043 ve Lentinus tigrinus OBCC 3007 miselleri taşıyıcı olarak kullanılan naylon temizlik süngeri üzerine immobilize edilmiştir. Fungusların immobilize misellerinin ayrı ayrı ve birlikte kültür edilmeleri durumundaki lakkaz ve mangan peroksidaz aktiviteleri karşılaştırılmıştır. Saf Pleurotus ostreatus OBCC 6043 ve Lentinus tigrinus OBCC 3007 kültürlerinin maksimum lakkaz aktiviteleri, sırası ile, 53.73 ve 27.58 U/L iken maksimum mangan peroksidaz aktiviteleri 12.54 ve 52.02 U/L olarak belirlenmiştir. Kokültür koşullarında lakkaz ve mangan peroksidaz aktiviteleri belirgin bir artış ile 319.28 ve 554.33 U/L seviyesine yükselmiştir. Pleurotus ostreatus OBCC 6043 ve Lentinus tigrinus OBCC 3007 birlikte üretildiklerinde lakkaz aktivitesi ayrı ayrı kültür edilmelerine göre, sırası ile,  5.94 ve  11.58 kat fazla bulunmuştur. Diğer taraftan mangan peroksidaz aktivitesi aynı sıra ile  44.21 ve  10.66 kat gibi belirgin biçimde arttırılabilmiştir. Sunulan birlikte kültür çalışmasının sonuçları, hem lakkaz hem de mangan peroksidaz için literatürde sunulan değerlerin çoğuna göre önemli derecede yüksektir.  

Anahtar Kelimeler

• İmmobilizasyon,Lakkaz,Lentinus tigrinus,Mangan peroksidaz,Pleurotus ostreatus.

Increasing of Laccase and Manganese Peroxidase Activity by Co-Culture of Immobilized Pleurotus ostreatus and Lentinus tigrinus Mycelia

Öz

In this study it has firstly reported the using of immobilized fungal mycelia in fungal co-culture studies to enhance activity of lignin-modifying enzymes. For this purpose, the mycelia of Pleurotus ostreatus OBCC 6043 and Lentinus tigrinus OBCC 3007 were immobilized on nylon scouring pad as carrier material. . The immobilized mycelia of the fungi were compared in terms of their laccase and manganese peroxidase activities in mono- and co-culture conditions. The maximum laccase activities of pure Pleurotus ostreatus OBCC 6043 and Lentinus tigrinus OBCC 3007 cultures were determined as 53.73 and 27.58 U/L, respectively, while the maximum manganese peroxidase activities were 12.54 and 52.02 U/L. In co-culture conditions, distinct enhancement was observed in laccase and manganese peroxidase activities as 319.28 and 554.33 U/L, respectively. In the case of laccase, enzyme activity was  5.94 and  11.58 times higher than that of Pleurotus ostreatus OBCC 6043 and Lentinus tigrinus OBCC 3007 mono-cultures, respectively. On the other hand, manganese peroxidase activity could be improved distinctly,  44.21 and  10.66 fold higher values than the corresponding ones. The results of the present co-culture study are significantly higher than most of the reported results in the literature, not only for laccase but also for manganese peroxidase activity.

Anahtar Kelimeler

• Immobilization,Laccase,Lentinus tigrinus,Manganese peroxidase,Pleurotus ostreatus

Kaynakça

1. Agnieszka K., Christian G., Marcel A., André F., Marius R., Thierry T,. LAC3, a new low redox potential laccase from Trametes sp. strain C30 obtained as a recombinant protein in yeast, Enzyme Microb. Technol., 36, 34-41, (2005).
2. Asiegbu F.O., Paterson A., Smith, J.E., The effects of co-fungal cultures and supplementation with carbohydrate adjuncts on lignin biodegradation and substrate digestibility, World J. Microbiol. Biotechnol., 12, 273–279, (1996).
3. Bader J., Mast-Gerlach E., Popović M.K., Bajpai R., Stahl U., Relevance of microbial coculture fermentations in biotechnology, J. Appl. Microbiol., 109(2), 371-387, (2010).
4. Baldrian P., Increase of laccase activity during interspecific interactions of white-rot fungi, FEMS Microbiol. Ecol., 50, 245-253, (2004).
5. Baldrian, P., Fungal laccases – occurrence and properties, FEMS Microbiol. Rev., 30, 215–242, (2006).
6. Chi Y., Hatakka A., Maijala P., Can co-culturing of two white-rot fungi increase lignin degradation and the production of lignin-degrading enzymes? Int. Biodeter. Biodeg., 59, 32–39, (2007).
7. Cupul W.C., Abarca G.H., Carrera D.M., Vázquez R.R., Enhancement of ligninolytic enzyme activities in a Trametes maxima–Paecilomyces carneus co-culture: Key factors revealed after screening using a Plackett–Burman experimental design, Electr. J. Biotechnol., 17, 114–121, (2014).
8. Dong Y.C., Wang W., Hu Z.C., Fu M.L., Chen Q.H., The synergistic effect on production of lignin-modifying enzymes through submerged co-cultivation of Phlebia radiata, Dichomitus squalens and Ceriporiopsis subvermispora using agricultural residues, Bioprocess Biosyst. Eng., 35, 751–760, (2012).

9. D’Souza D.T., Tiwari R., Sah A.K., Raghukumar C., Enhanced production of laccase by a marine fungus during treatment of colored effluents and synthesis dyes, Enzyme Microb. Technol., 38, 504–511, (2006).
10. Dwivedi P., Vivekanand V., Pareek N., Sharma A., Singh R.P., Co-cultivation of mutant Penicillium oxalicum SAUE-3.510 and Pleurotus ostreatus for simultaneous biosynthesis of xylanase and laccase under solid-state fermentation, New Biotechnol., 28 (6), 616-626, (2011).
11. Elisashvili V., Kachlishvili E., Physiological regulation of laccase and manganese peroxidase production by white-rot Basidiomycetes, J. Biotechnol., 144, 37-42, (2009).
12. Elisashvili V., Kachlishvili E., Penninckx M., Effect of growth substrate, method of fermentation, and nitrogen source on lignocellulose-degrading enzymes production by white-rot basidiomycetes, J. Ind. Microbiol. Biotechnol., 35, 1531–1538, (2008).
13. Fink-Boots M., Malarczyk E., Leonowicz A., Increased enzymatic activities and levels of superoxide anion and phenolic compounds in cultures of basidiomycetes after temperature stress, Acta Biotechnol., 19, 319–330, (1999).
14. Flores C., Casasanero R., Trejo-Hernandez M.R., Galindo E, Serrano-Carreo L., Production of laccases by Pleurotus ostreatus in submerged fermentation in co-culture with Trichoderma viride, J. Appl. Microbiol., 108, 810–817, (2010).
15. Hailei W., Guangli Y., Ping L., Yanchang G., Jun L., Guosheng L., Jianming Y., Overproduction of Trametes versicolor laccase by making glucose starvation using yeast, Enzyme Microbial Technol., 45, 146–149, (2009).
16. Hailei W., Chaozhi T., Guangli Y., Ping L., A novel membrane-surface liquid co-culture to improve the production of laccase from Ganoderma lucidum, Biochem. Eng. J., 80, 27– 36, (2013).
17. Hiscox J., Baldrian P., Rogers H.J., Boddy L., Changes in oxidative enzyme activity during interspecific mycelial interactions involving the white-rot fungus Trametes versicolor, Fungal Gen. Biol., 47, 562–571, (2010).
18. Hu H.L., van den Brink J., Gruben B.S., Wösten H.A.B., Gu J.D., de Vries R.P., Improved enzyme production by co-cultivation of Aspergillus niger and Aspergillus oryzae and with other fungi, Int. Biodeter. Biodeg., 65, 248-252, (2011).
19. Ijoma G.N., Tekere M., Potential microbial applications of co-cultures involving ligninolytic fungi in the bioremediation of recalcitrant xenobiotic compounds, Int. J. Environ. Sci. Technol., 14(8), 1787–1806, (2017).
20. Jaszek M., Grzywnowicz K., Malarczyk E., Leonowicz A., Enhanced extracellular laccase activity as a part of the response system of white rot fungi Trametes versicolor and Abortiporus biennis to paraquat-caused oxidative stress conditions, Pestic. Biochem. Physiol. 85, 147–154, (2006).
21. Jegatheesan M., Eyini M., Response Surface Methodology Mediated Modulation of Laccase Production by Polyporus arcularius, Arab. J. Sci. Eng., 40, 1809–1818, (2015).
22. Kiiskinen L.L., Saloheimo M., Molecular cloning and expression in Saccharomyces cerevisiae of a laccase gene from the ascomycete Melanocarpus albomyces, Appl. Environ. Microbiol., 70, 137–144, (2004).
23. Kiiskinen L.L., Rättö M., Kruus K., Screening for novel laccase-producing microbes, J. Appl. Microbiol., 97, 640–646, (2004).
24. Koroleva O.V., Stepanova E.V., Gavrilova V.P., Yakovleva N.S., Landesman E.O., Yavmetdinov I.S., Yaropolov A.I., Laccase and Mn-peroxidase production by Coriolus hirsutus strain 075 in a jar fermentor, J. Biosci. Bioeng., 93, 449-455, (2002).
25. Kuhar F., Castiglia V., Levin L., Enhancement of laccase production and malachite green decolorization by co-culturing Ganoderma lucidum and Trametes versicolor in solid-state fermentation, Int. Biodeter. Biodeg., 104, 238-243, (2015).
26. Kunamneni A., Plou F.J., Ballesteros A., Alcalde M., Laccases and their applications: a patent review, Recent Pat. Biotechnol., 2, 10–24, (2008).
27. Li P., Wang H.L., Liu G.S., Li X., Yao J.M., The effect of carbon source succession on laccase activity in the co-culture process of Ganoderma lucidum and a yeast, Enzyme Microb. Technol., 48, 1–6, (2011).
28. Machado K.M.G., Matheus D.R., Bononi V.L.R., Ligninolytic enzymes production and Remazol Brilliant Blue R decolorization by tropical brazilian basidiomycetes fungi, Brazilian J. Microbiol., 36, 246-252, (2005).
29. Marková E., Kotik M., Křenková A., Man P., Haudecoeur R., Boumendjel A., Hardré R., Mekmouche Y., Courvoisier-Dezord E., Réglier M., Martínková L., Recombinant tyrosinase from Polyporus arcularius: Overproduction in Escherichia coli, Characterization, and Use in a Study of Aurones as Tyrosinase Effectors, J. Agric. Food Chem.., 64(14), 2925-2931, (2016).
30. Miller G.L., Use of dinitrosalicylic acid reagent for determination of reducing sugar, Anal. Chem., 31, 426-428, (1959).
31. Niku-Paavola M., Raaska L., Itävaara M., Detection of white-rot fungi by a non-toxic stain, Mycol. Res., 94, 27-31, (1990).
32. Qi-he C., Krügener S., Hirth T., Rupp S., Zibek S., Co-cultured Production of Lignin-Modifying Enzymes with White-Rot Fungi, Appl. Biochem. Biotechnol., 165,700–718, (2011).
33. Okino L.K., Machado K.M.G., Fabris C., Bononi V.L.R., Ligninolytic activity of tropical rainforest basidiomycetes, World J. Microbiol. Biotechnol., 16, 889-893, (2000).
34. Rivela I., Rodriguez-Couto S., Sanroman A., Extracellular ligninolytic enzyme production by Phanerochaete chrysosporium in a new solid-state bioreactor, Biotechnol. Lett., 22, 1443-1447, (2000).
35. Rodriguez-Couto S., Production of laccase and decolouration of the textile dye Remazol Brilliant Blue R in temporary immersion bioreactors, J. Hazard. Mat., 194, 297–302, (2011).
36. Rodriguez-Couto S., Herrera, J.L.T., Industrial and biotechnological applications of laccases: a review, Biotechnol. Adv., 24, 500–513, (2006)
37. Searle P., The berthelot or indophenol reaction and its use in the analytical chemistry of nitrogen. A review, Analyst., 109, 549–568, (1984).
38. Singh N., Devi A., Jaryal R., Rani K., An Ecofriendly and Efficient Strategy for Cost Effective Production of Lignocellulotic Enzymes, Waste Biomass Valor., DOI 10.1007/s12649-017-9861-9, (2017).
39. Sun Y., Cheng J., Hydrolysis of lignocellulosic material from ethanol production: a review, Bioresour. Technol., 83, 1–11, (2002).
40. Theerachat M., Emond S., Cambon E., Bordes F., Marty A., Nicaud J.M., Chulalaksananukul W., Guieysse D., Remaud-Siméon M., More S., Engineering andproduction of laccase from Trametes versicolor in the yeast Yarrowia lipolytica, Bioresour. Technol., 125, 267–274, (2012).
41. Ürek R.Ö., Kasikara Pazarlıoğlu N., Purification and partial characterization of manganase peroxidase from immobilized Phanarochaete chrysosporium, Process Biochem., 39, 2061-2068, (2003)
42. Vinogradova S.P., Kushnir S.N., Biosynthesis of Hydrolytic Enzymes during Cocultivation of Macro- and Micromycetes, Appl. Biochem. Microbiol., 39(6), 573–575, (2003).
43. Wang H., Peng L., Ding Z. , Wu J., Shi G., Stimulated laccase production of Pleurotus ferulae JM301 fungus by Rhodotorula mucilaginosa yeast in co-culture, Process Biochem., 50, 901–905, (2015).