Abstract
Dictyoglomus turgidum is a hyperthermophilic, anaerobic, gram-negative bacterium that shows an array of putative glycoside hydrolases (GHs) encoded by its genome, a feature that makes this microorganism very interesting for biotechnological applications. The aim of this work is the characterization of a hyperthermophilic GH5, Dtur_0671, of D. turgidum, annotated as endoglucanase and herein named DturCelB in agreement to DturCelA, which was previously characterized. The synthetic gene was expressed in Escherichia coli. The purified recombinant enzyme is active as a monomer (40 kDa) and CD structural studies showed a conserved α/β structure at different temperatures (25 and 70 °C) and high thermoresistance (Tm of 88 °C). Interestingly, the enzyme showed high endo-β-1,4-mannanase activity vs various mannans, but low endo-β-1,4 glucanase activity towards carboxymethylcellulose. The K M and V max of DturCelB were determined for both glucomannan and CMC: they were 4.70 mg/ml and 473.1 μmol/min mg and 1.83 mg/ml and 1.349 μmol/min mg, respectively. Its optimal activity towards temperature and pH resulted to be 70 °C and pH 5.4, respectively. Further characterization highlighted good thermal stability (~ 50% of enzymatic activity after 2 h at 70 °C) and pH stability over a broad range (> 90% of activity after 1 h in buffer, ranging pH 5–9); resistance to chemicals was also observed.
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References
Acharya S, Chaudhary A (2012) Bioprospecting thermophiles for cellulase production: a review. Braz J Microbiol 43:844–856
Álvarez C, Reyes-Sosa FM, Díez B (2016) Enzymatic hydrolysis of biomass from wood. Microb Biotechnol 9:149–156
Aulitto M, Fusco S, Fiorentino G, Limauro D, Pedone E, Bartolucci S, Contursi P (2017) Thermus thermophilus as source of thermozymes for biotechnological applications: homologous expression and biochemical characterization of an α-galactosidase. Microb Cell Fact 16:28
Brumm PJ, Hermanson S, Luedtke J, Mead DA (2011a) Identification, cloning and characterization of Dictyoglomus turgidum CelA, an endoglucanase with cellulose and mannanase activity. J Life Sci 5:488–496
Brumm PJ, Hermanson S, Hochstein B, Boyum J, Hermersmann N, Gowda K, Mead DA (2011b) Mining Dictyoglomus turgidum for enzymatically active carbohydrases. Appl Biochem Biotechnol 163:205–214
Brumm PJ, Gowda K, Robb FT, Mead DA (2016) The complete genome sequence of hyperthermophile Dictyoglomus turgidum DSM 6724™ reveals a specialized carbohydrate fermentor. Front Microbiol 20:1979
Capolupo L, Faraco V (2016) Green methods of lignocellulose pretreatment for biorefinery development. Appl Microbiol Biotechnol 100:9451–9467
Chauhan PS, Puri N, Sharma P, Gupta N (2012) Mannanases: microbial sources, production, properties and potential biotechnological applications. Appl Microbiol Biotechnol 93:1817–1830
Chen CY, Huang YC, Yang TY, Jian JY, Chen WL, Yang CH (2016) Degradation of konjac glucomannan by Thermobifida fusca thermostable beta-mannanase from yeast transformant. Int J Biol Macromol 82:1–6
Chhabra S, Shockley K, Ward DE, Kelly RM (2002) Regulation of endo-acting glycosyl hydrolases in the hyperthermophilic bacterium Thermotoga maritima grown on glucan- and mannan-based polysaccharides. Appl Environ Microbiol 68:545–554
Contursi P, Fusco S, Limauro D, Fiorentino G (2013) Host and viral transcriptional regulators in Sulfolobus: an overview. Extremophiles 17:881–895
Del Giudice I, Limauro D, Pedone E, Bartolucci S, Fiorentino G (2013) A novel arsenate reductase from the bacterium Thermus thermophilus HB27: its role in arsenic detoxification. Biochim Biophys Acta 1834:2071–2079
Dhawan S, Kaur J (2007) Microbial mannanases: an overview of production and applications. J Crit Rev Biotechnol 27:197–216
Do BC, Dang TT, Berrin JG, Haltrich D, To KA, Sigoillot JC, Yamabhai M (2009) Cloning, expression in Pichia pastoris, and characterization of a thermostable GH5 mannan endo-1,4-beta-mannosidase from Aspergillus niger BK01. Microb Cell Fact 13:59
Dos Santos CR, Paiva JH, Meza AN, Cota J, Alvarez TM, Ruller R et al (2012) Molecular insights into substrate specificity and thermal stability of a bacterial GH5-CBM27 endo-1,4-β-d-mannanase. J Struct Biol 177:469–476
Elleuche S, Schröder C, Sahm K, Antranikian G (2014) Extremozymes–biocatalysts with unique properties from extremophilic microorganisms. Curr Opin Biotechnol 29:116–123
Elleuche S, Schäfers C, Blank S, Schröder C, Antranikian G (2015) Exploration of extremophiles for high temperature biotechnological processes. Curr Opin Microbiol 25:113–119
Gusakov AV, Kondratyeva EG, Sinitsyn AP (2011) Comparison of two methods for assaying reducing sugars in the determination of carbohydrase activities. Int J Anal Chem 2011:283658
Han Y, Dodd D, Hespen CW, Ohene-Adjei S, Schroeder CM, Mackie RI et al (2010) Comparative analyses of two thermophilic enzymes exhibiting both beta-1,4 mannosidic and beta-1,4 glucosidic cleavage activities from Caldanaerobius polysaccharolyticus. J Bacteriol 192:4111–4121
Kolpak FJ, Blackwell J (1976) Determination of the structure of cellulose II. Macromolecules 9:273–278
Kublanov IV, Perevalova AA, Slobodkina GB, Lebedinsky AV, Bidzhieva SK, Kolganova TV et al (2009) Biodiversity of thermophilic prokaryotes with hydrolytic activities in hot springs of Uzon Caldera, Kamchatka (Russia). Appl Environ Microbiol 75:286–291
Limauro D, Cannio R, Fiorentino G, Rossi M, Bartolucci S (2001) Identification and molecular characterization of an endoglucanase gene, celS, from the extremely thermophilic archaeon Sulfolobus solfataricus. Extremophiles 5:213–219
Limauro D, D’Ambrosio K, Langella E, De Simone G, Galdi I, Pedone C, Bartolucci S (2010) Exploring the catalytic mechanism of the first dimeric Bcp: functional, structural and docking analyses of Bcp4 from Sulfolobus solfataricus. Biochimie 92:1435–1444
Lombard V, Golaconda Ramulu H, Drula E, Coutinho PM, Henrissat B (2014) The carbohydrate-active enzymes database (CAZy) in 2013. Nucleic Acids Res 42:D490–D495
Malgas S, van Dyk JS, Pletshke BI (2015) A review of the enzymatic hydrolysis of mannans and synergic interactions between β-mannanase, β-mannosidase and α-galactosidase. Word J Microbiol Biotechnol 31:1167–1175
Mendoza NS, Arai M, Kawaguchi T, Yoshida T, Joson LM (1994) Purification and properties of mannanase from Bacillus subtilis. World J Microbiol Biotechnol 10:551–555
Nelson N (1944) A photometric adaptation of the Somogyi method for the determination of glucose. J Biol Chem 153:376–380
Pereira JH, Chen Z, McAndrew RP, Sapra R, Chhabra SR, Sale KL, Simmons BA, Adams PD (2010) Biochemical characterization and crystal structure of endoglucanase Cel5A from the hyperthermophilic Thermotoga maritima. J Struct Biol 172:372–379
Rytioja J, Hildén K, Yuzon J, Hatakka A, De Vries RP, Mäkelä MR (2014) Plant-polysaccharide-degrading enzymes from Basidiomycetes. Microbiol Mol Biol Rev 78:614–649
Sadhu S, Maiti TK (2013) Cellulase Production by Bacteria: a review. Brit Microbiol Res J 3:235–258
Sarcinelli C, Fiorentino G, Pizzo E, Bartolucci S, Limauro D (2016) Discovering antioxidant molecules in the archaea domain: peroxiredoxin Bcp1 from Sulfolobus solfataricus Protects H9c2 cardiomyoblasts from oxidative stress. Archaea 2016:7424870
Sarmiento F, Peralta R, Blamey JM (2015) Cold and hot extremozymes: industrial relevance and current trends. Front Bioeng Biotechnol 20:148
Scheller HV, Ulvskov P (2010) Hemicelluloses. Annu Rev Plant Biol 61:263–289
Seesom W, Thongket P, Yamamoto T, Takenaka S, Sakamoto T, Sukhumsirichart W (2017) Purification, characterization, and overexpression of an endo-1,4-β-mannanase from thermotolerant Bacillus sp. SWU60. World J Microbiol Biotechnol 33:53
Sreerama N, Woody RW (2000) Estimation of protein secondary structure from circular dichroism spectra: comparison of CONTIN, SELCON, and CDSSTR methods with an expanded reference set. Anal Biochem 287:252–260
Sreerama N, Venyaminov SY, Woody RW (1999) Estimation of the number of α-helical and β-strand segments in proteins using circular dichroism spectroscopy. Protein Sci 8:370–380
Sreerama N, Venyaminov SY, Woody RW (2000) Estimation of protein secondary structure from circular dichroism spectra: inclusion of denatured proteins with native proteins in the analysis. Anal Biochem 287:243–251
Srivastava PK, Kapoor M (2017) Production, properties, and applications of endo-β-mannanases. Biotechnol Adv 35:1–19
Svetlichny VA, Svetlichnaya TP (1988) Dictyoglomus turgidus sp. nov., a new extremely thermophilic eubacterium isolated from hot springs of the Uzon volcano caldera. Mikrobiologiia 57:364–369
Wilson B, Whelan K (2017) Prebiotic inulin-type fructans and galacto-oligosaccharides: definition, specificity, function, and application in gastrointestinal disorders. J Gastroenterol Hepatol 32:64–68
Xu BZ, Hagglund P, Stalbrand H, Janson JC (2002) Endo-beta-1,4-mannanases from blue mussel, Mytilus edulis: purification, characterization, and mode of action. J Biotechnol 92:267–277
Yamabhai M, Sak-Ubol S, Srila W, Haltrich D (2016) Mannan biotechnology: from biofuels to health. Crit Rev Biotechnol 36:32–42
Yang H, Shi P, Lu H, Wang H, Luo H, Huang H et al (2015) A thermophilic β-mannanase from Neosartorya fischeri P1 with broad pH stability and significant hydrolysis ability of various mannan polymers. Food Chem 15:283–289
Zakaria MM, Yamamoto S, Yagi T (1998) Purification and characterization of an endo-1,4-β-mannanase from Bacillus subtilis KU-1. FEMS Microbiol Lett 158:25–31
Zhang J, He ZM, Hu K (2000) Purification and characterization of beta-mannanase from Bacillus licheniformis for industrial use. Biotechnol Lett 22:1375–1378
Zhang M, Chen XL, Zhang ZH, Sun CY, Chen LL, He HL, Zhou BC, Zhang YZ (2009) Purification and functional characterization of endo-beta-mannanase MAN5 and its application in oligosaccharide production from konjac flour. Appl Microbiol Biotechnol 83:865–873
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This work was supported by BIOPOLIS: PON03PE_00107_1 CUP: E48C14000030005.
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Communicated by H. Atomi.
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Fusco, F.A., Ronca, R., Fiorentino, G. et al. Biochemical characterization of a thermostable endomannanase/endoglucanase from Dictyoglomus turgidum . Extremophiles 22, 131–140 (2018). https://doi.org/10.1007/s00792-017-0983-6
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DOI: https://doi.org/10.1007/s00792-017-0983-6