Abstract
Fossil fuels are a crucial resource for the global economy, but they also contribute to greenhouse gas emissions and environmental pollution. Lignocellulosic biomass, which includes cellulose, hemicellulose, and lignin obtained from plants, is a promising alternative to fossil fuels. It can help address these problems while reducing environmental impact. Enzymatic pre-treatment is used to degrade lignocellulosic biomass into subunits. The degradation of the hemicellulose structure involves accessory enzymes of industrial importance, such as α-glucuronidase. α-glucuronidases (EC 3.2.1.139) catalyze the hydrolysis of the α-1,2-glycosidic bond between α-D-glucuronic acid (GlcA) or its 4-o-methyl ether form (MeGlcA) and d-xylose units in the structure of xylooligosaccharides. The aim of this study was cloning, heterologous expression and biochemical characterization of the α-glucuronidase enzyme from the thermophilic bacterium Geobacillus kaustophilus. With this aim, the codon optimized α-glucuronidase gene was cloned into pQE-30 vector, overexpressed in E. coli BL21 (DE3), and purified with nickel affinity chromatography. The biochemical characterization of the purified α-glucuronidase revealed that the enzyme has activity at elevated temperatures between 65-90 °C. Additionally, Geobacillus kaustophilus α-glucuronidase enzyme showed higher activity at acidic pH values from pH 4.0 to 6.5. This is the first study to report the gene cloning, recombinant expression and biochemical characterization of α-glucuronidase which could be used as accessory enzyme from a thermophilic bacterium Geobacillus kaustophilus.
Keywords
Thermostable Geobacillus kaustophilus α-glucuronidase enzymes Lignocellulosic biomass Hemicellulolytic enzymes
References
- Aalbers F, Turkenburg JP, Davies GJ, Dijkhuizen L, Lammerts van Bueren A. 2015. Structural and functional characterization of a novel family GH115 4-O-methyl-α-glucuronidase with specificity for decorated arabinogalactans. J Mol Biol, 427(24): 3935-3946.
- Adıgüzel AO. 2013. Biyoetanolün genel özellikleri ve üretimi için gerekli hammadde kaynakları. Bitlis Eren Üniv Fen Bil Derg, 2(2): 204-220.
- Akkaya A, Ensari Y, Ozseker EE, Batur OO, Buyuran G, Evran S. 2023. Recombinant production and biochemical characterization of thermostable arabinofuranosidase from acidothermophilic alicyclobacillus acidocaldarius. Protein J, 42(4): 437-450.
- Arevalo-Gallegos A, Ahmad Z, Asgher M, Parra-Saldivar R, Iqbal HMN. 2017. Lignocellulose: A sustainable material to produce value-added products with a zero waste approach-A review. Int J Biol Macromol, 99: 308-318.
- Chimphango AFA, Görgens JF, van Zyl WH. 2016. In situ enzyme aided adsorption of soluble xylan biopolymers onto cellulosic material. Carbohydr Polym, 143: 172-178.
- Chong SL, Derba-Maceluch M, Koutaniemi S, Gómez LD, McQueen-Mason SJ, Tenkanen M, Mellerowicz EJ. 2015. Active fungal GH115 α-glucuronidase produced in Arabidopsis thaliana affects only the UX1-reactive glucuronate decorations on native glucuronoxylans. BMC Biotechnol, 15(1): 56.
- Dalia S, Gali G, Gil S, Yuval S. 2004. Effect of dimer dissociation on activity and thermostability of the α-glucuronidase from geobacillus stearothermophilus: Dissecting the different oligomeric forms of family 67 glycoside hydrolases. J Bacteriol, 186(20): 6928-6937.
- Demirjian DC, Morís-Varas F, Cassidy CS. 2001. Enzymes from extremophiles. Curr Opin Chem Biol, 5(2): 144-151.
- Ezeilo UR, Zakaria II, Huyop F, Wahab RA. 2017. Enzymatic breakdown of lignocellulosic biomass: the role of glycosyl hydrolases and lytic polysaccharide monooxygenases. Biotechnol. Biotechnol Equip, 31(4): 647-662.
- Jaramillo PMD, Gomes HAR, Monclaro AV, Silva COG, Filho EXF. 2015. Lignocellulose-degrading enzymes. Fungal Biomolec, 2015: 73-85.
- Lee CC, Kibblewhite RE, Wagschal K, Li R, Robertson GH, Orts WJ. 2012. Isolation and characterization of a novel GH67 α-glucuronidase from a mixed culture. J Ind Microbiol Biotechnol, 39(8): 1245-1251.
- Lee HV, Hamid SBA, Zain SK. 2014. Conversion of lignocellulosic biomass to nanocellulose: Structure and chemical process. Sci World J, 2014: 631013.
- Maitan-Alfenas GP, Visser EM, Guimarães VM. 2015. Enzymatic hydrolysis of lignocellulosic biomass: Converting food waste in valuable products. Curr Opin Food Sci, 1(1): 44-49.
- Mohapatra SB, Manoj N. 2019. Structure of an α-glucuronidase in complex with Co2+ and citrate provides insights into the mechanism and substrate recognition in the family 4 glycosyl hydrolases. Biochem Biophys Res Commun, 518(2): 197-203.
- Østby H, Hansen LD, Horn SJ, Eijsink VGH, Várnai A. 2020. Enzymatic processing of lignocellulosic biomass: principles, recent advances and perspectives. J Ind Microbiol Biotechnol, 47(9): 623-657.
- Rogowski A, Baslé A, Farinas CS, Solovyova A, Mortimer JC, Dupree P, Gilbert HJ, Bolam DN. 2014. Evidence that GH115 α-glucuronidase activity, which is required to degrade plant biomass, is dependent on conformational flexibility. J Biol Chem, 289(1): 53-64.
- Rosa L, Ravanal MC, Mardones W, Eyzaguirre J. 2013. Characterization of a recombinant α-glucuronidase from Aspergillus fumigatus. Fungal Biol, 117(5): 380-387.
- Septiningrum K, Ohi H, Waeonukul R, Pason P, Tachaapaikoon C, Ratanakhanokchai K, Sermsathanaswadi J, Deng L, Prawitwong P, Kosugi A. 2015. The GH67 α-glucuronidase of Paenibacillus curdlanolyticus B-6 removes hexenuronic acid groups and facilitates biodegradation of the model xylooligosaccharide hexenuronosyl xylotriose. Enzyme Microb Technol, 71: 28-35.
- Shao W, Obi S, Puls J, Wiegel J. 1995. Purification and Characterization of the (alpha)-Glucuronidase from Thermoanaerobacterium sp. Strain JW/SL-YS485, an Important Enzyme for the Utilization of Substituted Xylans. Appl Environ Microbiol, 61(3): 1077-1081.
- Suresh C, Kitaoka M, Hayashi K. 2003. A thermostable non-xylanolytic α-glucuronidase of Thermotoga maritima MSB8. Biosci Biotechnol Biochem, 67(11): 2359-2364.
- Turner P, Mamo G, Karlsson EN. 2007. Potential and utilization of thermophiles and thermostable enzymes in biorefining. Microb Cell Fact, 6(1): 9.
- Van den Burg B. 2003. Extremophiles as a source for novel enzymes. Curr Opin Microbiol, 6(3): 213-218.
- Wang F, Ouyang D, Zhou Z, Page SJ, Liu D, Zhao X. 2021. Lignocellulosic biomass as sustainable feedstock and materials for power generation and energy storage. J Energy Chem, 57: 247-280.
- Wang W, Yan R, Nocek BP, Vuong TV, Di Leo R, Xu X, Cui H, Gatenholm P, Toriz G, Tenkanen M, Savchenko A, Master ER. 2016. Biochemical and structural characterization of a five-domain GH115 α-glucuronidase from the marine bacterium Saccharophagus degradans 2-40T*. J Biol Chem, 291(27): 14120-14133.
- Yan R, Vuong TV, Wang W, Master ER. 2017. Action of a GH115 α-glucuronidase from Amphibacillus xylanus at alkaline condition promotes release of 4-O-methylglucopyranosyluronic acid from glucuronoxylan and arabinoglucuronoxylan. Enzyme Microb Technol, 104: 22-28.
- Yeoman CJ, Han Y, Dodd D, Schroeder CM, Mackie RI, Cann IK. 2010. Chapter 1 - Thermostable Enzymes as Biocatalysts in the Biofuel Industry. In Advances in Applied Microbiology, Academic Press, London, UK, pp: 1-55.
- Zaide G, Shallom D, Shulami S, Zolotnitsky G, Golan G, Baasov T, Shoham G, Shoham Y. 2001. Biochemical characterization and identification of catalytic residues in alpha-glucuronidase from Bacillus stearothermophilus T-6. Eur. J Biochem, 268(10): 3006-3016.
Abstract
Fossil fuels are a crucial resource for the global economy, but they also contribute to greenhouse gas emissions and environmental pollution. Lignocellulosic biomass, which includes cellulose, hemicellulose, and lignin obtained from plants, is a promising alternative to fossil fuels. It can help address these problems while reducing environmental impact. Enzymatic pre-treatment is used to degrade lignocellulosic biomass into subunits. The degradation of the hemicellulose structure involves accessory enzymes of industrial importance, such as α-glucuronidase. α-glucuronidases (EC 3.2.1.139) catalyze the hydrolysis of the α-1,2-glycosidic bond between α-D-glucuronic acid (GlcA) or its 4-o-methyl ether form (MeGlcA) and d-xylose units in the structure of xylooligosaccharides. The aim of this study was cloning, heterologous expression and biochemical characterization of the α-glucuronidase enzyme from the thermophilic bacterium Geobacillus kaustophilus. With this aim, the codon optimized α-glucuronidase gene was cloned into pQE-30 vector, overexpressed in E. coli BL21 (DE3), and purified with nickel affinity chromatography. The biochemical characterization of the purified α-glucuronidase revealed that the enzyme has activity at elevated temperatures between 65-90 °C. Additionally, Geobacillus kaustophilus α-glucuronidase enzyme showed higher activity at acidic pH values from pH 4.0 to 6.5. This is the first study to report the gene cloning, recombinant expression and biochemical characterization of α-glucuronidase which could be used as accessory enzyme from a thermophilic bacterium Geobacillus kaustophilus.
Keywords
Thermostable Geobacillus kaustophilus α-glucuronidase enzymes Lignocellulosic biomass Hemicellulolytic enzymes
References
- Aalbers F, Turkenburg JP, Davies GJ, Dijkhuizen L, Lammerts van Bueren A. 2015. Structural and functional characterization of a novel family GH115 4-O-methyl-α-glucuronidase with specificity for decorated arabinogalactans. J Mol Biol, 427(24): 3935-3946.
- Adıgüzel AO. 2013. Biyoetanolün genel özellikleri ve üretimi için gerekli hammadde kaynakları. Bitlis Eren Üniv Fen Bil Derg, 2(2): 204-220.
- Akkaya A, Ensari Y, Ozseker EE, Batur OO, Buyuran G, Evran S. 2023. Recombinant production and biochemical characterization of thermostable arabinofuranosidase from acidothermophilic alicyclobacillus acidocaldarius. Protein J, 42(4): 437-450.
- Arevalo-Gallegos A, Ahmad Z, Asgher M, Parra-Saldivar R, Iqbal HMN. 2017. Lignocellulose: A sustainable material to produce value-added products with a zero waste approach-A review. Int J Biol Macromol, 99: 308-318.
- Chimphango AFA, Görgens JF, van Zyl WH. 2016. In situ enzyme aided adsorption of soluble xylan biopolymers onto cellulosic material. Carbohydr Polym, 143: 172-178.
- Chong SL, Derba-Maceluch M, Koutaniemi S, Gómez LD, McQueen-Mason SJ, Tenkanen M, Mellerowicz EJ. 2015. Active fungal GH115 α-glucuronidase produced in Arabidopsis thaliana affects only the UX1-reactive glucuronate decorations on native glucuronoxylans. BMC Biotechnol, 15(1): 56.
- Dalia S, Gali G, Gil S, Yuval S. 2004. Effect of dimer dissociation on activity and thermostability of the α-glucuronidase from geobacillus stearothermophilus: Dissecting the different oligomeric forms of family 67 glycoside hydrolases. J Bacteriol, 186(20): 6928-6937.
- Demirjian DC, Morís-Varas F, Cassidy CS. 2001. Enzymes from extremophiles. Curr Opin Chem Biol, 5(2): 144-151.
- Ezeilo UR, Zakaria II, Huyop F, Wahab RA. 2017. Enzymatic breakdown of lignocellulosic biomass: the role of glycosyl hydrolases and lytic polysaccharide monooxygenases. Biotechnol. Biotechnol Equip, 31(4): 647-662.
- Jaramillo PMD, Gomes HAR, Monclaro AV, Silva COG, Filho EXF. 2015. Lignocellulose-degrading enzymes. Fungal Biomolec, 2015: 73-85.
- Lee CC, Kibblewhite RE, Wagschal K, Li R, Robertson GH, Orts WJ. 2012. Isolation and characterization of a novel GH67 α-glucuronidase from a mixed culture. J Ind Microbiol Biotechnol, 39(8): 1245-1251.
- Lee HV, Hamid SBA, Zain SK. 2014. Conversion of lignocellulosic biomass to nanocellulose: Structure and chemical process. Sci World J, 2014: 631013.
- Maitan-Alfenas GP, Visser EM, Guimarães VM. 2015. Enzymatic hydrolysis of lignocellulosic biomass: Converting food waste in valuable products. Curr Opin Food Sci, 1(1): 44-49.
- Mohapatra SB, Manoj N. 2019. Structure of an α-glucuronidase in complex with Co2+ and citrate provides insights into the mechanism and substrate recognition in the family 4 glycosyl hydrolases. Biochem Biophys Res Commun, 518(2): 197-203.
- Østby H, Hansen LD, Horn SJ, Eijsink VGH, Várnai A. 2020. Enzymatic processing of lignocellulosic biomass: principles, recent advances and perspectives. J Ind Microbiol Biotechnol, 47(9): 623-657.
- Rogowski A, Baslé A, Farinas CS, Solovyova A, Mortimer JC, Dupree P, Gilbert HJ, Bolam DN. 2014. Evidence that GH115 α-glucuronidase activity, which is required to degrade plant biomass, is dependent on conformational flexibility. J Biol Chem, 289(1): 53-64.
- Rosa L, Ravanal MC, Mardones W, Eyzaguirre J. 2013. Characterization of a recombinant α-glucuronidase from Aspergillus fumigatus. Fungal Biol, 117(5): 380-387.
- Septiningrum K, Ohi H, Waeonukul R, Pason P, Tachaapaikoon C, Ratanakhanokchai K, Sermsathanaswadi J, Deng L, Prawitwong P, Kosugi A. 2015. The GH67 α-glucuronidase of Paenibacillus curdlanolyticus B-6 removes hexenuronic acid groups and facilitates biodegradation of the model xylooligosaccharide hexenuronosyl xylotriose. Enzyme Microb Technol, 71: 28-35.
- Shao W, Obi S, Puls J, Wiegel J. 1995. Purification and Characterization of the (alpha)-Glucuronidase from Thermoanaerobacterium sp. Strain JW/SL-YS485, an Important Enzyme for the Utilization of Substituted Xylans. Appl Environ Microbiol, 61(3): 1077-1081.
- Suresh C, Kitaoka M, Hayashi K. 2003. A thermostable non-xylanolytic α-glucuronidase of Thermotoga maritima MSB8. Biosci Biotechnol Biochem, 67(11): 2359-2364.
- Turner P, Mamo G, Karlsson EN. 2007. Potential and utilization of thermophiles and thermostable enzymes in biorefining. Microb Cell Fact, 6(1): 9.
- Van den Burg B. 2003. Extremophiles as a source for novel enzymes. Curr Opin Microbiol, 6(3): 213-218.
- Wang F, Ouyang D, Zhou Z, Page SJ, Liu D, Zhao X. 2021. Lignocellulosic biomass as sustainable feedstock and materials for power generation and energy storage. J Energy Chem, 57: 247-280.
- Wang W, Yan R, Nocek BP, Vuong TV, Di Leo R, Xu X, Cui H, Gatenholm P, Toriz G, Tenkanen M, Savchenko A, Master ER. 2016. Biochemical and structural characterization of a five-domain GH115 α-glucuronidase from the marine bacterium Saccharophagus degradans 2-40T*. J Biol Chem, 291(27): 14120-14133.
- Yan R, Vuong TV, Wang W, Master ER. 2017. Action of a GH115 α-glucuronidase from Amphibacillus xylanus at alkaline condition promotes release of 4-O-methylglucopyranosyluronic acid from glucuronoxylan and arabinoglucuronoxylan. Enzyme Microb Technol, 104: 22-28.
- Yeoman CJ, Han Y, Dodd D, Schroeder CM, Mackie RI, Cann IK. 2010. Chapter 1 - Thermostable Enzymes as Biocatalysts in the Biofuel Industry. In Advances in Applied Microbiology, Academic Press, London, UK, pp: 1-55.
- Zaide G, Shallom D, Shulami S, Zolotnitsky G, Golan G, Baasov T, Shoham G, Shoham Y. 2001. Biochemical characterization and identification of catalytic residues in alpha-glucuronidase from Bacillus stearothermophilus T-6. Eur. J Biochem, 268(10): 3006-3016.
Details
| Primary Language | English |
|---|---|
| Subjects | Biocatalysis and Enzyme Technology |
| Journal Section | Research Articles |
| Authors | |
| Early Pub Date | February 17, 2024 |
| Publication Date | March 15, 2024 |
| Submission Date | December 19, 2023 |
| Acceptance Date | January 15, 2024 |
| Published in Issue | Year 2024 Volume: 7 Issue: 2 |
