Improvement of Catalytic Efficiency, Thermo-stability and Dye Decolorization Capability of Pleurotus ostreatus IBL-02 laccase by Hydrophobic Sol Gel Entrapment

Chemistry Central Journal

Muhammad Asgher¹  Shagufta Kamal¹  Hafiz Muhammad Nasir Iqbal¹ 

[1] Industrial Biotechnology Laboratory, Department of Chemistry and Biochemistry, University of Agriculture, Faisalabad, Pakistan

关键词: Decolorization;    Waste water effluent;    Textile dye;    Kinetics;    Sol–gel immobilization;    PAGE;    Laccase;    P. ostreatus IBL-02;   

Others  :  788066
DOI  :  10.1186/1752-153X-6-110

 received in 2012-08-07, accepted in 2012-09-27,  发布年份 2012

【 摘 要 】

Background

In serious consideration of the worldwide environmental issues associated with the extensive use of the textile dyes and effluents generated thereof, the scientists across the world are in search for potential treatment technologies for their treatment. In such scenario the ligninolytic enzymes provide a potential alternative because they are cost effective, eco-friendly and can be applied to wide range of dye containing industrial effluents.

Results

Laccase produced from Pleurotus ostreatus IBL-02 during decolorization of the reactive textile dye Drimarene brilliant red K-4BL (DBR K-4BL) was purified and immobilized by hydrophobic gel entrapment. The crude laccase was 4.2-fold purified with specific activity of 573.52 U/mg after passing through the DEAE-Sepharose ion exchange and Sephadex-G-100 chromatography columns. P. ostreatus IBL-02 laccase was found to be a homogenous monomeric protein as evident by single band corresponding to 67 kDa on native and sodium dodesylsulfate polyacrylamide gel electrophoresis (PAGE). The laccase was immobilized by entrapment in Sol–gel matrix of trimethoxysilane (T) and proplytetramethoxysilane (P) prepared using different T:P molar ratios. The free and immobilized laccases were compared to investigate the effect of immobilization on catalytic efficiency and thermo-stability features. Laccase immobilized in the Sol–gel of 1:5 T:P ratio was optimally active and thermo-stable fraction at pH 5, 60°C with half-life of 3 h and 50 min. Laccases immobilized in 1:2 and 1:5 T:P ratio gels had significantly higher K_m (83 and100mM) and V_max (1000 and 1111 mM/mg) values as compared to free laccase. After 5 h reaction time varying decolorization percentages with a maximum of 100% were achieved for different dyes and effluents.

Conclusions

In summary, P. ostreatus IBL-02 laccase was immobilized by entrapping in a Sol–gel matrix with an objective to enhance its catalytic and stability properties. Sol–gel entrapped laccase presented potential efficiency as a biocatalyst when applied for decolorization of different dyes and effluents. The main benefits of the Sol–gel matrix immobilization processes are the eco-friendly approach, chemical free and energy saving reaction conditions.

【 授权许可】

   
2012 Asgher et al.; licensee Chemistry Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

Figure 8.

Figure 9.

Figure 10.

【 参考文献 】

• [1]Wesenberg D, Kyriakides I, Agathos SN: White-rot fungi and their enzymes for the treatment of industrial dye effluents. Biotechnol Adv 2003, 22:161-187.
• [2]Colao MC, Lupino S, Garzillo AM, Buonocore V, Ruzzi M: Heterologous expression of lcc1 gene from Trametes trogii in Pichia pastoris and characterization of the recombinant enzyme. Microb Cell Fact 2006, 5:31. BioMed Central Full Text
• [3]Asgher M, Bhatti HN, Ashraf M, Legge RL: Recent developments in biodegradation of industrial pollutants by white rot fungi and their enzyme system. Biodegradation 2008, 19:771-783.
• [4]Kim J-M, Park S-M, Kim D-H: Heterologous expression of a tannic acid-inducible laccase3 of Cryphonectria parasitica in Saccharomyces cerevisiae. BMC Biotechnol 2010, 10:18. BioMed Central Full Text
• [5]Stoilova I, Krastanov A, Stanchev V: Properties of crude laccase from Trametes versicolor produced by solid-substrate fermentation. Adv Biosci Biotechnol 2010, 1:208-215.
• [6]Asgher M, Iqbal HMN: Characterization of a novel manganese peroxidase purified from solid state culture of Trametes versicolor IBL-04. BioRes 2011, 6:4317-4330.
• [7]Reiss R, Ihssen J, Thöny-Meyer L: Bacillus pumilus laccase: a heat stable enzyme with a wide substrate spectrum. BMC Biotechnol 2011, 11:9. BioMed Central Full Text
• [8]Asgher M, Iqbal HMN, Asad MJ: Kinetic characterization of purified laccase produced from Trametes versicolor IBL-04 in solid state bio-processing of corncobs. BioRes 2012, 7:1171-1188.
• [9]Kunamneni A, Camarero S, García-Burgos C, Plou FJ, Ballesteros A, Alcalde M: Engineering and Applications of fungal laccases for organic synthesis. Microb Cell Fact 2008, 7:32. BioMed Central Full Text
• [10]Asgher M, Iqbal HMN, Irshad M: Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of corncobs. BMC Biotechnol 2012, 12:46. BioMed Central Full Text
• [11]Cheng J, Randall A, Baldi M: Prediction of Protein Stability Changes for Single-Site Mutations Using Support Vector Machines. Prot Str Func Bioinf 2006, 62:1125-1132.
• [12]Almeida VM, Branco CRC, Assis SA, Vieira IJC, Braz-Filho R, Branco A: Synthesis of naringin 6"-ricinoleate using immobilized lipase. Chem Central J 2012, 6:41. BioMed Central Full Text
• [13]Iqbal HMN, Asgher M: Characterization and decolorization applicability of xerogel matrix immobilized manganese peroxidase produced from Trametes versicolor IBL-04. Protein Pept Lett 2012. In-Press, PPL-EPUB-20120925-3
• [14]Irshad M, Bahadur BA, Anwar Z, Yaqoob M, Ijaz A, Iqbal HMN: Decolorization applicability of sol–gel matrix-immobilized laccase produced from Ganoderma leucidum using agro-industrial waste. BioRes 2012, 7(3):4249-4261.
• [15]Saratale RG, Saratale GD, Chang JS, Govindwar SP: Outlook of bacterial decolorization and degradation of azo dyes: a review. J Taiwan Inst Chem Eng 2011, 42:138-157.
• [16]Asgher M, Asad MJ, Bhatti HN, Legge RL: Hyperactivation and thermo-stabilization of Phanerochaete chrysosporium lignin peroxidase by immobilization in xerogels. World J Microbiol Biotechnol 2007, 23:525-531.
• [17]Kamal S, Asgher M, Khalil-ur-Rehman , Zahir ZA: Hyperproduction of laccase by Pleurotus ostreatus IBL-02 during decolorization of drimarene brilliant red K-4BL. Fresen Environ Bull 2011, 20:1478-1486.
• [18]Chen S, Ge W, Buswell JA: Biochemical and molecular characterization of a laccase from the edible straw mushroom Volvariella volvacea. Eur J Biochem 2004, 271:318-328.
• [19]Mansur M, Arias ME, Patino JLC, Gonzalez MFAE: The white-rot fungus Pleurotus ostreatus secretes laccase isozymes with different substrate specificities. Mycologia 2003, 95:1013-1020.
• [20]Miao L, Zhang G, Wang H, Ng T: Purification and Characterization of a Laccase from the edible wild mushroom Tricholoma mongolicum. J Microbiol Biotechnol 2010, 20:1069-1076.
• [21]Nagai M, Sato T, Saito K, Kawata M: Purification and characterization of an extracellular laccase from the edible mushroom Lentinula edodes, and decolorization of chemically different dyes. Appl Microbiol Biotechnol 2002, 60:327-335.
• [22]Du Z, Sun XB: Purification and characterization of laccase from Curvularia trifol. Adv Mat Res 2010, 116:2215-2219.
• [23]Perez J, Rubia TD, Hamman OB, Martinez J: Phanerochaete flavidoalba laccase induction and modification of manganese peroxidase is enzyme pattern in decolorized olive oil mill wastewaters. Appl Environ Microbiol 1998, 64:2722-2729.
• [24]Qiu H, Xu C, Huang X, Ding Y, Qu Y, Gao P: Immobilization of laccase on nanoporous gold: comparative studies on the immobilization strategies and the particle size effects. J Phys Chem 2009, 113:2521-2525.
• [25]Clifford JS, Legge RL: Use of water to evaluate hydrophobicity of organically-modified Xerogel enzyme supports. Biotechnol Bioeng 2005, 92:231-237.
• [26]Rekuc A, Bryjak J, Szymanska K, Jarzebski AB: Laccase immobilization on mesostructured cellular foams afford preparations with ultra-high activity. Proc Biochem 2009, 44:191-198.
• [27]Huang J, Liu Y, Wang X: Silanized palygorskite for lipase immobilization. J Mol Catal B: Enz 2009, 57:10-15.
• [28]Arica M, Altıntas B, Bayramoglu G: Immobilization of laccase onto spacer-arm attached non-porous poly(GMA/EGDMA) beads: Application for textile dye degradation. Biores Technol 2009, 100:665-669.
• [29]Singh G, Bhalla A, Capalash N, Sharma P: Characterization of immobilized laccase from γ-proteobacterium JB: Approach towards the development of biosensor for the detection of phenolic compounds. Indian J Sci Technol 2010, 2:48-53.
• [30]Prasad KK, Mohan SV, Bhaskar YV, Ramanaiah SV, Babu VL, Pati BR, Sarma PN: Laccase production using Pleurotus ostreatus 1804 immobilized on PUF cubes in batch and packed bed reactors: influence of culture conditions. J Microbiol 2005, 43:301-307.
• [31]Wang F, Guo C, Yang L, Liu CZ: Magnetic mesoporous silica nanoparticles: Fabrication and their laccase immobilization performance. Biores Technol 2010, 101:8931-8935.
• [32]Pazarlioglu NK, Sariisik M, Telefoncu A: Laccase production by Trametes versicolor and application to denim washing. Proc Biochem 2005, 40:1673-1678.
• [33]Bayramoglu G, Yilmaz M, Arica MY: Reversible immobilization of laccase to poly(4-vinylpyridine) grafted and Cu(II) chelated magnetic beads: Biodegradation of reactive dyes. Biores Technol 2010, 101:6615-6621.
• [34]Maas R, Chaudhari S: Adsorption and biological decolorization of azo dye reactive red 2 in semicontinuous anaerobic reactors. Proc Biochem 2005, 40:699-705.
• [35]Bradford MM: A rapid and sensitive method for quantification of microgram quantities of protein utilizing the principle of protein dye binding. Anal Biochem 1976, 72:248-254.
• [36]Laemmli UK: Cleavage of structural proteins during assembly of head of bacteriophage T4. Nature 1970, 227:680-685.