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Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs

BACKGROUND: Cost-effective production of industrially important enzymes is a key for their successful exploitation on industrial scale. Keeping in view the extensive industrial applications of lignin peroxidase (LiP), this study was performed to purify and characterize the LiP from an indigenous str...

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Autores principales: Asgher, Muhammad, Iqbal, Hafiz Muhammad Nasir, Irshad, Muhammad
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3442999/
https://www.ncbi.nlm.nih.gov/pubmed/22862820
http://dx.doi.org/10.1186/1472-6750-12-46
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author Asgher, Muhammad
Iqbal, Hafiz Muhammad Nasir
Irshad, Muhammad
author_facet Asgher, Muhammad
Iqbal, Hafiz Muhammad Nasir
Irshad, Muhammad
author_sort Asgher, Muhammad
collection PubMed
description BACKGROUND: Cost-effective production of industrially important enzymes is a key for their successful exploitation on industrial scale. Keeping in view the extensive industrial applications of lignin peroxidase (LiP), this study was performed to purify and characterize the LiP from an indigenous strain of Trametes versicolor IBL-04. Xerogel matrix enzyme immobilization technique was applied to improve the kinetic and thermo-stability characteristics of LiP to fulfil the requirements of the modern enzyme consumer sector of biotechnology. RESULTS: A novel LiP was isolated from an indigenous T. versicolor IBL-04 strain. T. versicolor IBL-04 was cultured in solid state fermentation (SSF) medium of corn cobs and maximum LiP activity of 592 ± 6 U/mL was recorded after five days of incubation under optimum culture conditions. The crude LiP was 3.3-fold purified with specific activity of 553 U/mg after passing through the DEAE-cellulose and Sephadex-G-100 chromatography columns. The purified LiP exhibited a relatively low molecular weight (30 kDa) homogenous single band on native and SDS-PAGE. The LiP was immobilized by entrapping in xerogel matrix of trimethoxysilane (TMOS) and proplytetramethoxysilane (PTMS) and maximum immobilization efficiency of 88.6% was achieved. The free and immobilized LiPs were characterized and the results showed that the free and immobilized LiPs had optimum pH 6 and 5 while optimum temperatures were 60°C and 80°C, respectively. Immobilization was found to enhance the activity and thermo-stability potential of LiP significantly and immobilized LiP remained stable over broad pH and temperature range as compare to free enzyme. Kinetic constants K(m) and V(max) were 70 and 56 μM and 588 and 417 U/mg for the free and immobilized LiPs, respectively. Activity of this novel extra thermo-stable LiP was stimulated to variable extents by Cu(2+), Mn(2+) and Fe(2+) whereas, Cystein, EDTA and Ag(+) showed inhibitory effects. CONCLUSIONS: The indigenously isolated white rot fungal strain T. versicolor IBL-04 showed tremendous potential for LiP synthesis in SSF of corncobs in high titters (592 U/mL) than other reported Trametes (Coriolus, Polyporus) species. The results obtained after dual phase characterization suggested xerogel matrix entrapment a promising tool for enzyme immobilization, hyper-activation and stabilization against high temperature and inactivating agents. The pH and temperature optima, extra thermo-stability features and kinetic characteristics of this novel LiP of T. versicolor IBL-04 make it a versatile enzyme for various industrial and biotechnological applications.
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spelling pubmed-34429992012-09-15 Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs Asgher, Muhammad Iqbal, Hafiz Muhammad Nasir Irshad, Muhammad BMC Biotechnol Research Article BACKGROUND: Cost-effective production of industrially important enzymes is a key for their successful exploitation on industrial scale. Keeping in view the extensive industrial applications of lignin peroxidase (LiP), this study was performed to purify and characterize the LiP from an indigenous strain of Trametes versicolor IBL-04. Xerogel matrix enzyme immobilization technique was applied to improve the kinetic and thermo-stability characteristics of LiP to fulfil the requirements of the modern enzyme consumer sector of biotechnology. RESULTS: A novel LiP was isolated from an indigenous T. versicolor IBL-04 strain. T. versicolor IBL-04 was cultured in solid state fermentation (SSF) medium of corn cobs and maximum LiP activity of 592 ± 6 U/mL was recorded after five days of incubation under optimum culture conditions. The crude LiP was 3.3-fold purified with specific activity of 553 U/mg after passing through the DEAE-cellulose and Sephadex-G-100 chromatography columns. The purified LiP exhibited a relatively low molecular weight (30 kDa) homogenous single band on native and SDS-PAGE. The LiP was immobilized by entrapping in xerogel matrix of trimethoxysilane (TMOS) and proplytetramethoxysilane (PTMS) and maximum immobilization efficiency of 88.6% was achieved. The free and immobilized LiPs were characterized and the results showed that the free and immobilized LiPs had optimum pH 6 and 5 while optimum temperatures were 60°C and 80°C, respectively. Immobilization was found to enhance the activity and thermo-stability potential of LiP significantly and immobilized LiP remained stable over broad pH and temperature range as compare to free enzyme. Kinetic constants K(m) and V(max) were 70 and 56 μM and 588 and 417 U/mg for the free and immobilized LiPs, respectively. Activity of this novel extra thermo-stable LiP was stimulated to variable extents by Cu(2+), Mn(2+) and Fe(2+) whereas, Cystein, EDTA and Ag(+) showed inhibitory effects. CONCLUSIONS: The indigenously isolated white rot fungal strain T. versicolor IBL-04 showed tremendous potential for LiP synthesis in SSF of corncobs in high titters (592 U/mL) than other reported Trametes (Coriolus, Polyporus) species. The results obtained after dual phase characterization suggested xerogel matrix entrapment a promising tool for enzyme immobilization, hyper-activation and stabilization against high temperature and inactivating agents. The pH and temperature optima, extra thermo-stability features and kinetic characteristics of this novel LiP of T. versicolor IBL-04 make it a versatile enzyme for various industrial and biotechnological applications. BioMed Central 2012-08-03 /pmc/articles/PMC3442999/ /pubmed/22862820 http://dx.doi.org/10.1186/1472-6750-12-46 Text en Copyright ©2012 Asgher et al.; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Article
Asgher, Muhammad
Iqbal, Hafiz Muhammad Nasir
Irshad, Muhammad
Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs
title Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs
title_full Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs
title_fullStr Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs
title_full_unstemmed Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs
title_short Characterization of purified and Xerogel immobilized Novel Lignin Peroxidase produced from Trametes versicolor IBL-04 using solid state medium of Corncobs
title_sort characterization of purified and xerogel immobilized novel lignin peroxidase produced from trametes versicolor ibl-04 using solid state medium of corncobs
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3442999/
https://www.ncbi.nlm.nih.gov/pubmed/22862820
http://dx.doi.org/10.1186/1472-6750-12-46
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