Isolation and partial purification of fungal ligninolytic enzymes from the forest soil fungi isolated from Bhadra Wildlife Sanctuary

Abstract

Screening was done for the isolation of effective lignin degraders from the forest soil samples, by providing lignin as a carbon source through the enrichment method, which leads to the isolation of 8 effective fungal isolates among 14 isolates. Submerged fermentation was done for the production of ligninolytic enzymes with the effective microorganisms by providing Guiaicol as a carbon source. The assay of laccase, lignin peroxidise activity and specific activity was done after the incubation intervals of 2, 4, 6, 7, 8, 10 and 12 days at 27±2°C under shake culture condition. Partially purified protein content was estimated by using Lowry’s method. Pleurotus sp. and Phanerochaetae chrysosporium are more effective at the 2nd and 7th days of incubation for the production of laccase and lignin peroxidases among the effective isolates.

Keywords

laccase / lignin peroxidase / extracellular enzyme / fungi / forest soil

References

[1] Abdel-Raheem A M, Ali E H (2004). Lignocellulolytic enzyme production by aquatic hyphomycetes species isolated from the Nile’s delta region. Mycopathologia, 157(3): 277-286

[2] Akin D E, Rigsby L L, Sethuraman A, Morrison W H 3rd, Gamble G R, Eriksson K E L (1995). Alterations in structure, chemistry, and biodegradability of grass lignocellulose treated with the white rot fungi Ceriporiopsis subvermispora and Cyathus stercoreus. Appl Environ Microbiol, 61(4): 1591-1598

[3] Arora D S, Chander M, Gill P K (2002). Involvement of lignin peroxidase, manganese peroxidase and laccase in the degradation and selective ligninolysis of wheat straw. Int Bioterior Biodegrad, 50: 115-120

[4] Bajpai P (2004). Biological bleaching of chemical pulps. Crit Rev Biotechnol, 24(1): 1-58

[5] Baldrian P, Gabriel J (2003). Lignocellulose degradation by Pleurotus ostreatus in the presence of cadmium. FEMS Microbiol Lett, 220(2): 235-240

[6] Bergmeyer H U (1974). Methods of Enzymatic Analysis 1, 2nd Ed, New York: Academic Press, 1-495

[7] Bosco F, Ruggeri B, Sassi G (1999). Performances of a trickle bed reactor (TBR) for exoenzyme production by Phanerochaete chrysosporium: influence of a superficial liquid velocity. Chem Eng Sci, 54(15-16): 3163-3169

[8] Carlsen M, Nielsen J (2001). Influence of carbon source on α-amylase production by Aspergillus oryzae. Appl Microbiol Biotechnol, 57(3): 346-349

[9] Champion H G, Seth S K (1968). A Revised Survey of the forest types of India. New Delhi: Govt. of India Press, 1-404

[10] Coll P M, Fernández-Abalos J M, Villanueva J R, Santamaría R, Pérez P (1993). Purification and characterization of a phenoloxidase (laccase) from the lignin-degrading basidiomycete PM1 (CECT 2971). Appl Environ Microbiol, 59(8): 2607-2613

[11] Das N, Sengupta S, Mukherjee M (1997). Importance of laccase in vegetative growth of pleurotus Florida. Appl Environ Microbiol, 63(10): 4120-4122

[12] Dhakar K, Pandey A (2013). Laccase Production from a Temperature and pH Tolerant Fungal Strain of Trametes hirsuta (MTCC 11397). Enzyme Res, 2013: 869062Available at:

[13] Domsch K H, Gams W (1972). Fungi in Agricultural Soils. London: Longmans Green, 1-290

[14] Dritsa V, Rigas F, Natsis K, Marchant R (2007). Characterization of a fungal strain isolated from a polyphenol polluted site. Bioresour Technol, 98(9): 1741-1747

[15] Eggert C, Temp U, Dean J F, Eriksson K E (1996a). A fungal metabolite mediates degradation of non-phenolic lignin structures and synthetic lignin by laccase. FEBS Lett, 391(1-2): 144-148

[16] Eggert C, Temp U, Eriksson K E L (1996b). The ligninolytic system of the white rot fungus Pycnoporus cinnabarinus: purification and characterization of the laccase. Appl Environ Microbiol, 62(4): 1151-1158

[17] Eisenlord S D, Freedman Z, Zak D R, Xue K, He Z, Zhou J (2013). Microbial mechanisms mediating increased soil C storage under elevated atmospheric N deposition. Appl Environ Microbiol, 79(4): 1191-1199

[18] Ellis M B (1971). Dematiaceous Hyphomycetes, Kew: Commonwealth Mycological Institute. England, 1-608

[19] Ellis M B (1976). More Dematiaceous Hyphomycetes, Kew: Commonwealth Mycological Institute. England, 1-507

[20] Ellis M B, Ellis J P (1997). Microfungi on Land Plants: An Identification Handbook. London: Croom Helm, Richmond Publishers, 1-868

[21] Falcon M A, Rodriguez A, Carnicero A, Regalado V, Perestelo F, Milstein O, Fuente G L (1995). Isolation of microorganisms with lignin transformation potential from soil of Tenerife Island. Soil Biol Biochem, 27(2): 121-126

[22] Giardina P, Aurilia V, Cannio R, Marzullo L, Amoresano A, Siciliano R, Pucci P, Sannia G (1996). The gene, protein and glycan structures of laccase from Pleurotus ostreatus. Eur J Biochem, 235(3): 508-515

[23] Gilman J C (2001). A Manual of Soil Fungi, 2nd ed., <PublisherLocation><?Pub Caret?>New Delhi</PublisherLocation>: Biotech Books, 1-392

[24] Gochev V K, Krastanov A I (2007). Fungal Laccases. Bulg J Agric Sci, 13: 75-83

[25] Gold M H, Alic M (1993). Molecular biology of the lignin-degrading basidiomycete Phanerochaete chrysosporium. Microbiol Rev, 57(3): 605-622

[26] Hara T, Lim J Y, Fujio Y, Ueda S (1984). Purification and some properties of exopolygalacturonase from Aspergillus niger cultured in the medium containing Satsuna mandarin peel. 4th Conference on Recent Technologies in Agriculture, 31: 581-586

[27] Howard R L, Abotsi E, Rensburg E L J V, Howard S (2003). Lignocellulose biotechnology: issues of bioconversion and enzyme production. Afr J Biotechnol, 2: 602-619

[28] Kerem Z, Friesem D, Hadar Y (1992). Lignocellulose Degradation during Solid-State Fermentation: Pleurotus ostreatus versus Phanerochaete chrysosporium. Appl Environ Microbiol, 58(4): 1121-1127

[29] Lowry O H, Rosebrough N J, Farr A L, Randall R J (1951). Protein measurement with the folin phenol regent. J Gen Microbiol, 131: 3017-3027

[30] Lundell T, Hatakka A (1994). Participation of Mn(II) in the catalysis of laccase, manganese peroxidase and lignin peroxidase from Phelbia radiata. FEBS Lett, 348(3): 291-296

[31] Metuku R P, Burra S, Nidadavolu, Bindu S V S S S L H, Pabba S, Singaracharya M A (2011). Selection of highest lignolytic white rot fungus and its molecular identification. J Cell Tissue Research, 11: 2557-2562

[32] Moilanen A M, Lundell T, Vares T, Hatakka A (1996). A. Hatakka, Manganese and malonate are individual regulators for the production of lignin and manganese peroxidase isozymes and in the degradation of lignin by Phlebia radiate. Appl Microbiol Biotechnol, 45(6): 792-799

[33] Muñoz C, Guillén F, Martínez A T, Martínez M J (1997). Laccase isoenzymes of Pleurotus eryngii: characterization, catalytic properties, and participation in activation of molecular oxygen and Mn²⁺ oxidation. Appl Environ Microbiol, 63(6): 2166-2174

[34] Nagamani A, Kunwar I K, Manoharachary C (2006). Handbook of Soil Fungi. New Delhi: I. K. International Pvt. Ltd, 1-477

[35] Novotny C, Svobodova K, Erbanova P, Cajthaml T, Kasinath A, Lang E, Sasek V (2004). Ligninolytic fungi in bioremediation: extracellular enzyme production and degradation rate. Soil Biol Biochem, 36(10): 1545-1551

[36] Patil N P, Chaudhari B L (2010). Production and purification of pectinase by soil isolate Penicillium sp. and search for better Agro-residue for its SSF. Rec Res Sci Technol, 2: 36-42

[37] Périé F H, Reddy G V, Blackburn N J, Gold M H (1998). Purification and characterization of laccases from the white-rot basidiomycete Dichomitus squalens. Arch Biochem Biophys, 353(2): 349-355

[38] Piontek K, Smith A T, Blodig W (2001). Lignin peroxidase structure and function. Biochem Soc Trans, 29(Pt 2): 111-116

[39] Pitt J I (1979). The Genus Penicillium and its telomorphic states Eupenicillium and Talaromyces.London: Academic Press Inc Ltd, 1-634

[40] Pointing S B, Jones E B G, Vrijmoed L L P (2000). Optimization of laccase production by Pycnoporus sanguineus in submerged liquid culture. Mycologia, 92(1): 139-144

[41] Pozdnyakova N N (2012). Involvement of the ligninolytic system of white-rot and litter-decomposing fungi in the degradation of polycyclic aromatic hydrocarbons. Biotechnol Res Int, 2012: 243217

[42] Ruggeri B, Sassi G (2003). Experimental sensitivity analysis of a trickle bed bioreactor for lignin peroxidases production by Phanerochaetae chrysosporium. Process Biochem, 38(12): 1-8

[43] Sarkanen S, Razal R A, Piccariello T, Yamamoto E, Lewis N G (1991). Lignin peroxidase: toward a clarification of its role in vivo. J Biol Chem, 266(6): 3636-3643

[44] Tanaka H, Itakura S, Enoki A (1999). Hydroxyl radical generation by an extracellular low-molecular-weight substance and phenol oxidase activity during wood degradation by the white-rot basidiomycete Trametes versicolor. J Biotechnol, 75(1): 57-70

[45] Thurston C F (1994). The structure and function of fungal laccases. Microbiology, 140(1): 19-26

[46] Tien M, Kirk T K (1988). Lignin peroxidase of Phanerochate chrysosporium. Methods Enzymol, 161: 238-249

[47] Vicuna R (1988). Bacterial degradation of lignin. Enzyme Microb Technol, 10(11): 646-655

[48] Wu J, Xiao Y Z, Yu H Q (2005). Degradation of lignin in pulp mill wastewaters by white-rot fungi on biofilm. Bioresour Technol, 96(12): 1357-1363

[49] Xavier A M R B, Tavares A P M, Ferreira R, Amado F (2007). Trametes versicolor growth and laccase induction with by-products of pulp and paper industry. Electron J Biotechnol, 10(3): 444-451

[50] Yu Z, Zeng G M, Chen Y N, Zhang J C, Yu Y, Li H, Liu Z F, Tang L (2011). Effects of inoculation with Phanerochaete chrysosporium on remediation of pentachlorophenol-contaminated soil waste by composting. Process Biochem, 46(6): 1285-1291

[51] Zadrazil F, Gonser A, Lang E (1999). Influence of incubation temperature on the secretion of extracellulare lignolytic enzymes of Pleurotus and Dichomitus squalus into soil. Granada, Spain: Proceedings of the Conference on Enzymes in the environment

[52] Zimmermann W (1990). Degradation of lignin by bacteria. J Biotechnol, 13(2-3): 119-130

Acknowledgements

Authors are grateful to the Kuvempu University, Jnanasahyadri, Shankaraghatta, Karnataka, India, for providing necessary facilities and support for the completion of this work. They are indebted to their family and friends for their good wishes for the success of this work.

Compliance with ethics guidelines

Shivakuamar P. BANAKAR and B. THIPPESWAMY declare that they have no conflict of interest in the publication. This article does not contain any studies with human or animal subjects performed by any of the authors.

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