Skip to main content
Log in

Properties of an alkali-thermo stable xylanase from Geobacillus thermodenitrificans A333 and applicability in xylooligosaccharides generation

  • Original Paper
  • Published:
World Journal of Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

An extracellular thermo-alkali-stable and cellulase-free xylanase from Geobacillus thermodenitrificans A333 was purified to homogeneity by ion exchange and size exclusion chromatography. Its molecular mass was 44 kDa as estimated in native and denaturing conditions by gel filtration and SDS-PAGE analysis, respectively. The xylanase (GtXyn) exhibited maximum activity at 70 °C and pH 7.5. It was stable over broad ranges of temperature and pH retaining 88 % of activity at 60 °C and up to 97 % in the pH range 7.5–10.0 after 24 h. Moreover, the enzyme was active up to 3.0 M sodium chloride concentration, exhibiting at that value 70 % residual activity after 1 h. The presence of other metal ions did not affect the activity with the sole exceptions of K+ that showed a stimulating effect, and Fe2+, Co2+ and Hg2+, which inhibited the enzyme. The xylanase was activated by non-ionic surfactants and was stable in organic solvents remaining fully active over 24 h of incubation in 40 % ethanol at 25 °C. Furthermore, the enzyme was resistant to most of the neutral and alkaline proteases tested. The enzyme was active only on xylan, showing no marked preference towards xylans from different origins. The hydrolysis of beechwood xylan and agriculture-based biomass materials yielded xylooligosaccharides with a polymerization degree ranging from 2 to 6 units and xylobiose and xylotriose as main products. These properties indicate G. thermodenitrificans A333 xylanase as a promising candidate for several biotechnological applications, such as xylooligosaccharides preparation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Anand A, Kumar V, Satyanarayana T (2013) Characteristics of thermostable endoxylanase and β-xylosidase of the extremely thermophilic bacterium Geobacillus thermodenitrificans TSAA1 and its applicability in generating xylooligosaccharides and xylose from agro-residues. Extremophiles 17:357–366. doi:10.1007/s00792-013-0524-x

    Article  CAS  Google Scholar 

  • Archana A, Satyanarayana T (1997) Xylanase production by thermophilic Bacillus licheniformis A99 in solid state fermentation. Enzyme Microb Technol 21:12–17. doi:10.1016/S0141-0229(96)00207-4

    Article  CAS  Google Scholar 

  • Bastawde KB (1992) Xylan structure, microbial xylanases, and their mode of action. World J Microbiol Biotechnol 8:353–368. doi:10.1007/BF01198746

    Article  CAS  Google Scholar 

  • Biely P, Mislovicová D, Toman R (1985) Soluble chromogenic substrates for the assay of endo-1,4-beta-xylanases and endo-1,4-beta-glucanases. Anal Biochem 144(1):142–146. doi:10.1016/0003-2697(85)90095-8

    Article  CAS  Google Scholar 

  • Biely P, Hirsch J, la Grange DC, Van Zyl WH, Prior BA (2000) A chromogenic substrate for β-xylosidase coupled assay of α-glucuronidase. Anal Biochem 286:289–294. doi:10.1006/abio.2000.4810

    Article  CAS  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  Google Scholar 

  • Canakci S, Inan K, Kacagan M, Belduz AO (2007) Evaluation of arabinofuranosidase and xylanase activities of Geobacillus spp. isolated from some hot springs in Turkey. J Microbiol Biotechnol 17(8):1262–1270

    CAS  Google Scholar 

  • Canakci S, Cevher Z, Inan K, Tokgoz M, Bahar F, Kacagan M, Sal FA, Belduz AO (2012) Cloning, purification and characterization of an alkali-stable endoxylanase from thermophilic Geobacillus sp. 71. World J Microbiol Biotechnol 28:1981–1988. doi:10.1007/s11274-011-1000-3

    Article  CAS  Google Scholar 

  • Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, Henrissat B (2009) The Carbohydrate-Active EnZymes database (CAZy): an expert resource for glycogenomic. Nucleic Acids Res 37:D233–D238. doi:10.1093/nar/gkn663

    Article  CAS  Google Scholar 

  • Coleri A, Cokmus C, Ozcan B, Akkoc N, Akcelik M (2009) Isolation of alpha-glucosidase-producing thermophilic bacilli from hot springs of Turkey. Mikrobiologiia 78:68–78

    CAS  Google Scholar 

  • Collins T, Gerday C, Feller G (2005) Xylanases, xylanase families and extremophilic xylanases. FEMS Microbiol Rev 29:3–23. doi:10.1016/j.femsre.2004.06.005

    Article  CAS  Google Scholar 

  • Feng L, Wang W, Cheng J, Ren Y, Zhao G, Gao C, Tang Y, Liu X, Han W, Peng X, Liu R, Wang L (2007) Genome and proteome of long-chain alkane degrading Geobacillus thermodenitrificans NG80-2 isolated from a deep-subsurface oil reservoir. Proc Natl Acad Sci USA 104(13):5602–5607. doi:10.1073/pnas.0609650104

    Article  CAS  Google Scholar 

  • Fontes CM, Hall J, Hirst BH, Hazlewood GP, Gilbert HJ (1995) The resistance of cellulases and xylanases to proteolytic inactivation. Appl Microbiol Biotechnol 43:52–57. doi:10.1007/s002530050369

    Article  CAS  Google Scholar 

  • Gerasimova J, Kuisiene N (2012) Characterization of the novel xylanase from the thermophilic Geobacillus thermodenitrificans JK11. Mikrobiologiia 81(4):418–424. doi:10.1134/S0026261712040066

    CAS  Google Scholar 

  • Gessesse A (1998) Purification and properties of two thermostable alkaline xylanases from an alkaliphilic Bacillus sp. Appl Environ Microbiol 64:3533–3535

    CAS  Google Scholar 

  • Guo B, Chen XL, Sun CY, Zhou BC, Zhang YZ (2009) Gene cloning, expression and characterization of a new cold-active and salt-tolerant endo-β-1,4-xylanase from marine Glaciecola mesophila KMM 241. Appl Microbiol Biotechnol 84:1107–1115. doi:10.1007/s00253-009-2056-y

    Article  CAS  Google Scholar 

  • Juturu V, Wu JC (2012) Microbial xylanases: engineering, production and industrial applications. Biotechnol Adv 30(6):1219–1227. doi:10.1016/j.biotechadv.2011.11.006

    Article  CAS  Google Scholar 

  • Kaya F, Heitmann JA, Joyce TW (1995) Influence of surfactants on the enzymatic hydrolysis of xylan and cellulose. Tappi J 78:150–157

    CAS  Google Scholar 

  • Khandeparker R, Verma P, Deobagkar D (2011) A novel halotolerant xylanase from marine isolate Bacillus subtilis cho40: gene cloning and sequencing. New Biotechnol 28(6):814–821. doi:10.1016/j.nbt.2011.08.001

    Article  CAS  Google Scholar 

  • Khasin A, Alchanati I, Shoham Y (1993) Purification and characterization of a thermostable xylanase from Bacillus stearothermophilus T-6. Appl Environ Microbiol 59(6):1725–1730

    CAS  Google Scholar 

  • Krasuska E, Cardonica C, Tenorio JL, Testa G, Scordia D (2010) Potential land availability for energy crops production in Europe. Biofuels, Bioprod Biorefin 4:658–673. doi:10.1002/bbb.259

    Article  CAS  Google Scholar 

  • Kulkarni N, Shendye A, Rao M (1999) Molecular and biotechnological aspects of xylanases. FEMS Microbiol Rev 23(4):411–456. doi:10.1111/j.1574-6976.1999.tb00407.x

    Article  CAS  Google Scholar 

  • Kumar V, Satyanarayana T (2011) Applicability of thermo-alkali-stable and cellulase-free xylanase from a novel thermo-halo-alkaliphilic Bacillus halodurans in producing xylooligosaccharides. Biotechnol Lett 33:2279–2285. doi:10.1007/s10529-011-0698-1

    Article  CAS  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T-4. Nature 227:680–685. doi:10.1038/227680a0

    Article  CAS  Google Scholar 

  • Li N, Yang P, Wang Y, Luo H, Meng K, Wu N, Fan Y, Yao B (2008) Cloning, expression, and characterization of protease-resistant xylanase from Streptomyces fradiae var. k11. J Microbiol Biotechnol 18:410–416

    CAS  Google Scholar 

  • Liu B, Zhang N, Zhao C, Lin B, Xie L, Huang Y (2012) Characterization of a recombinant thermostable xylanase from hot spring thermophilic Geobacillus sp. TC-W7. J Microbiol Biotechnol 22(10):1388–1394. doi:10.4014/jmb.1203.03045

    Article  CAS  Google Scholar 

  • Mamo G, Hatti-Kaul R, Mattiasson B (2007) Fusion of carbohydrate binding modules from Thermotoga neapolitana with a family 10 xylanase from Bacillus halodurans S7. Extremophiles 11:169–177. doi:10.1007/s00792-006-0023-4

    Article  CAS  Google Scholar 

  • Matsudaira P (1987) Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem 262(21):10035–10038

    CAS  Google Scholar 

  • Maurelli L, Ionata E, La Cara F, Morana A (2013) Chestnut shell as unexploited source of fermentable sugars: effect of different pretreatment methods on enzymatic saccharification. Appl Biochem Biotechnol 170:1104–1118. doi:10.1007/s12010-013-0264-5

    Article  CAS  Google Scholar 

  • Menon G, Mody K, Keshri J, Jha B (2010) Isolation, purification, and characterization of haloalkaline xylanase from a marine Bacillus pumilus strain, GESF-1. Biotechnol Bioprocess Eng 15:998–1005. doi:10.1007/s12257-010-0116-x

    Article  CAS  Google Scholar 

  • Morrison D, van Dyk JS, Pletschke BI (2011) The effect of alcohols, lignin and phenolic compounds on the enzyme activity of Clostridium cellulovorans XynA. BioResources 6(3):3132–3141

    CAS  Google Scholar 

  • Moure A, Gullon P, Dominguez H, Parajo JC (2006) Advances in the manufacture, purification and applications of xylooligosaccharides as food additives and nutraceuticals. Process Biochem 41:1913–1923. doi:10.1016/j.procbio.2006.05.011

    Article  CAS  Google Scholar 

  • Nelson N (1944) A photometric adaptation of the Somogyi method for the determination of glucose. J Biol Chem 153:375–380

    CAS  Google Scholar 

  • Nielsen H, Brunak S, von Heijne G (1999) Machine learning approaches for the prediction of signal peptides and other protein sorting signals. Protein Eng 12:3–9. doi:10.1093/protein/12.1.3

    Article  CAS  Google Scholar 

  • Sato Y, Fukuda H, Zhou Y, Mikami S (2010) Contribution of ethanol-tolerant xylanase G2 from Aspergillus oryzae on Japanese sake brewing. J Biosci Bioeng 110:679–683. doi:10.1016/j.jbiosc.2010.07.015

    Article  CAS  Google Scholar 

  • Satyanarayana T, Sharma A, Mehta D, Puri AK, Kumar V, Nisha M, Joshi S (2012) Biotechnological applications of biocatalysts from the firmicutes Bacillus and Geobacillus species. In: Satyanarayana T, Johri BN, Prakash A (eds) Microorganisms in sustainable agriculture and biotechnology, part 2. Springer, Dordrecht, pp 343–379

    Chapter  Google Scholar 

  • Scordia D, Cosentino SL, Lee JW, Jeffries TW (2011) Dilute oxalic acid pretreatment for biorefining giant reed (Arundo donax L.). Biomass Bioenergy 35:3018–3024. doi:10.1016/j.biombioe.2011.03.046

    Article  CAS  Google Scholar 

  • Sellek GA, Chaudhuri JB (1999) Biocatalysis in organic media using enzymes from extremophiles. Enzyme Microb Technol 25:471–482. doi:10.1016/S0141-0229(99)00075-7

    Article  CAS  Google Scholar 

  • Sharma A, Adhikari S, Satyanarayana T (2007) Alkali-thermostable and cellulasefree xylanase production by an extreme thermophile Geobacillus thermoleovorans. World J Microbiol Biotechnol 23:483–490. doi:10.1007/s11274-006-9250-1

    Article  CAS  Google Scholar 

  • Subramaniyan S, Prema P (2000) Cellulase-free xylanases from Bacillus and other microorganisms. FEMS Microbiol Lett 183:1–7. doi:10.1111/j.1574-6968.2000.tb08925.x

    Article  CAS  Google Scholar 

  • Tan SS, Li DY, Jiang ZQ, Zhu YP, Shi B, Li LT (2008) Production of xylobiose form the autohydrolysis explosion liquor of corncob using Thermotoga maritima xylanase B (XynB) immobilized on nickel-chelated Eupergit C. Bioresour Technol 99:200–204. doi:10.1016/j.biortech.2006.12.005

    Article  CAS  Google Scholar 

  • Timell TE (1967) Recent progress in the chemistry of wood hemicelluloses. Wood Sci Technol 1:45–70. doi:10.1007/BF00592255

    Article  CAS  Google Scholar 

  • Tseng MJ, Yap MN, Ratanakhanokchai K, Kyu KL, Chen ST (2002) Purification and characterization of two cellulase free xylanases from an alkaliphilic Bacillus firmus. Enzyme Microb Technol 30:590–595. doi:10.1016/S0141-0229(02)00018-2

    Article  CAS  Google Scholar 

  • Vazquez MJ, Alonso JL, Domınguez H, Parajo JC (2000) Xylooligosaccharides: manufacture and applications. Trend Food Sci Technol 11:387–393. doi:10.1016/s0924-2244(01)00031-0

    Article  CAS  Google Scholar 

  • Verma D, Satyanarayana T (2012) Cloning, expression and applicability of thermo-alkali-stable xylanase of Geobacillus thermoleovorans in generating xylooligosaccharides from agro-residues. Bioresour Technol 107:333–338. doi:10.1016/j.biortech.2011.12.055

    Article  CAS  Google Scholar 

  • Vieille C, Zeikus GJ (2001) Hyperthermophilic enzymes: sources, uses, and molecular mechanisms for thermostability. Microbiol Mol Biol Rev 65(1):1–43. doi:10.1128/MMBR.65.1.1-43.2001

    Article  CAS  Google Scholar 

  • Viikari L, Kantelinen A, Sundquist J, Linko M (1994) Xylanases in bleaching: from an idea to the industry. FEMS Microbiol Rev 13:335–350. doi:10.1111/j.1574-6976.1994.tb00053.x

    Article  CAS  Google Scholar 

  • Woldesenbet F, Gupta N, Sharma P (2012) Statistical optimization of the production of a cellulase-free, thermo-alkali-stable, salt- and solvent-tolerant xylanase from Bacillus halodurans by solid state fermentation. Arch Appl Sci Res 4(1):524–535

    CAS  Google Scholar 

  • Wu S, Liu B, Zhang X (2006) Characterization of a recombinant thermostable xylanase from deep-sea thermophilic Geobacillus sp. MT-1 East Pacific. Appl Microbiol Biotechnol 72:1210–1216. doi:10.1007/s00253-006-0416-4

    Article  CAS  Google Scholar 

  • Yang R, Xu S, Wang Z, Yang W (2005) Aqueous extraction of corncob xylan and production of xylooligosaccharides. Food Sci Technol LEB 38:677–682. doi:10.1016/j.lwt.2004.07.023

    Article  CAS  Google Scholar 

  • Zhou J, Gao Y, Dong Y, Tang X, Li J, Xu B, Mu Y, Wu Q, Huang Z (2012) A novel xylanase with tolerance to ethanol, salt, protease, SDS, heat, and alkali from actinomycete Lechevalieria sp. HJ3. J Ind Microbiol Biotechnol 39:965–975. doi:10.1007/s10295-012-1113-1

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The authors would like to thank Dr. Luisa Maurelli and Dr. Daniela Landino for the skillful technical assistance, and Mr. Vito Carratore from the Protein Sequencing Core Facility at the Institute of Biosciences and Bioresources of the National Research Council in Naples, Italy, for the enzyme N-terminal sequence determination.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Francesco La Cara.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Marcolongo, L., La Cara, F., Morana, A. et al. Properties of an alkali-thermo stable xylanase from Geobacillus thermodenitrificans A333 and applicability in xylooligosaccharides generation. World J Microbiol Biotechnol 31, 633–648 (2015). https://doi.org/10.1007/s11274-015-1818-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11274-015-1818-1

Keywords

Navigation