Skip to main content
SpringerLink
Log in

Cloning, purification, and characterization of a thermostable α-l-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari

  • Biotechnologically Relevant Enzymes and Proteins
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

The gene, AbfAC26Sari, encoding an α-l-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari, was isolated, cloned, sequenced, and characterizated. On the basis of amino acid sequence similarities, this 57-kDa enzyme could be assigned to family 51 of the glycosyl hydrolase classification system. Characterization of the purified recombinant α-l-arabinofuranosidase produced in Escherichia coli BL21 revealed that it is active at a broad pH range (pH 4.5 to 9.0) and at a broad temperature range (45–85°C) and it has an optimum pH of 5.5 and an optimum temperature of 65°C. Kinetic experiment at 65°C with p-nitrophenyl α-l-arabinofuranoside as a substrate gave a V max and K m values of 1,019 U/mg and 0.139 mM, respectively. The enzyme had no apparent requirement of metal ions for activity, and its activity was strongly inhibited by 1 mM Cu2+ and Hg2+. The recombinant arabinofuranosidase released l-arabinose from arabinan, arabinoxylan, oat spelt xylan, arabinobiose, arabinotriose, arabinotetraose, and arabinopentaose. Endoarabinanase activity was not detected. These findings suggest that AbfAC26Sari is an exo-acting enzyme.

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

Access this article

Log in via an institution

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

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Aristidou A, Penttila M (2000) Metabolic engineering applications to renewable resource utilization. Curr Opin Biotechnol 11:187–198

    Article  CAS  Google Scholar 

  • Belduz AO, Lee EJ, Harman JG (1993) Mutagenesis of the cyclic AMP receptor protein of Escherichia coli: targeting positions 72 and 82 of the cyclic nucleotide binding pocket. Nuc Acids Res 21:1827–1835

    Article  CAS  Google Scholar 

  • Bezalel L, Shoham Y, Rosenberg E (1993) Characterization and delignification activity of a thermostable α-l-arabinofuranosidase from Bacillus stearothermophilus. Appl Microbiol Biotechnol 40:57–62

    Article  CAS  Google Scholar 

  • Brice RE, Morrison JM (1982) The degradation of isolated hemicelluloses and lignin-hemicellulose complexes by cell-free rumen hemicellulases. Carbohydr Res 101:93–100

    Article  CAS  Google Scholar 

  • Canakci S, Belduz AO, Saha BC, Yasar A, Ayaz FA, Yayli N (2007) Purification and characterization of a highly thermostable α-l-arabinofuranosidase from Geobacillus caldoxylolyticus TK4. Appl Microbiol Biotechnol 75:813–820

    Article  CAS  Google Scholar 

  • Degrassi G, Vindigni A, Venturi VA (2003) Thermostable α-l-arabinofuranosidase from xylonolytic Bacillus pumilus: purification and characterization. J Biotechnol 101:69–79

    Article  CAS  Google Scholar 

  • Emanuelsson O, Brunak S, Heijne G, Nielsen H (2007) Locating proteins in the cell using TargetP, SignalP and related tools. Nature Protocols 2:953–971

    Article  CAS  Google Scholar 

  • Gilead S, Shoham Y (1995) Purification and characterization of α-l-arabinofuranosidase from Bacillus stearothermophilus T-6. Appl Environ Microbiol 61:170–174

    Article  CAS  Google Scholar 

  • Gunata ZY, Brillouet JM, Voirin S, Baume R, Cordonnier R (1990) Purification and some properties of an α-l-arabinofuranosidase from Aspergillus niger. Action on grape monoterpenyl arabinofuranosylglucosides. J Agric Food Chem 38:772–776

    Article  CAS  Google Scholar 

  • Hata K, Tanaka M, Tsumuraya Y, Hashimoto Y (1992) α-l-Arabinofuranosidase from radish (Raphanus sativus L.) seeds. Plant Physiol 100:388–396

    Article  CAS  Google Scholar 

  • Hespell RB, O’Bryan PJ (1992) Purification and characterization of an α-l-arabinofuranosidase from Butyrivibrio fibrisolvens GS113. Appl Environ Microbiol 58:1082–1088

    Article  CAS  Google Scholar 

  • Hövel K, Shallom D, Niefind K, Belakhov V, Shoham G, Baasov T, Shoham Y, Schomburg D (2003) Crystal structure and snapshots along the reaction pathway of a family 51 α-l-Arabinofuranosidase. The EMBO Journal 22:4922–4932

    Article  Google Scholar 

  • Kaji A, Tagawa K, Ichimi T (1969) Properties of purified α-l-arabinofuranosidase from Aspergillus niger. Biochim Biophys Acta 171:186–188

    Article  CAS  Google Scholar 

  • Margolles A, de los Reyes-Gavilan CG (2003) Purification of a novel of α-l-arabinofuranosidase from Bifidobacterium longum B667. Appl Environ Microbiol 69:5096–5103

    Article  CAS  Google Scholar 

  • Matuso N, Kaneko S, Kuno A, Kobayashi H, Kusakabes I (2000) Purification, characterization and gene cloning of two α-l-arabinofuranosidases from Streptomyces chartreusis GS901. Biochem J 346:9–15

    Article  Google Scholar 

  • Miller GL (1959) Use of dinitrosalicyclic acid reagent for determination of reducing sugars. Anal Chem 31:426–428

    Article  CAS  Google Scholar 

  • Pikuta E, Lysenko A, Chuvilskaya N, Mendrock U, Hippe H, Suzina N, Nikitin D, Osipov G, Laurinavichius K (2000) Anoxybacillus pushchinensis gen. nov., sp. nov., a novel anaerobic, alkaliphilic, moderately thermophilic bacterium from manure, and description of Anoxybacillus flavithermus comb. nov. Int J Syst Evol Microbiol 50:2109–2117

    Article  CAS  Google Scholar 

  • Saha BC (2000) α-l-arabinofuranosidase: biochemistry, molecular biology and application in biotechnology. Biotechnol Adv 18:403–423

    Article  CAS  Google Scholar 

  • Saha BC (2003) Hemicellulose bioconversion. J Ind Microbiol Biotechnol 30:279–291

    Article  CAS  Google Scholar 

  • Seri K, Sanai K, Matsuo N, Kawakubo K, Xue C, Inoue S (1996) Arabinose selectively inhibits intestinal sucrase in uncompetitive manner and reduces glycemic response after sucrose ingestion in animals. Metabolism 45:1368–1374

    Article  CAS  Google Scholar 

  • Shallom D, Belakhovb V, Solomonb D, Gilead-Groppera S, Baasovb T, Shoham G, Shoham Y (2002a) The identification of acid-base catalyst of α-l-arabinofuranosidase from Geobacillus stearothermophilus T-6, a family 51 glycoside hydrolase. FEBS Letters 514:163–167

    Article  CAS  Google Scholar 

  • Shallom D, Belakhov V, Solomon D, Shoham G, Baasov T, Shoham Y (2002b) Detailed kinetic analysis and identification of the nucleophile in α-l-arabinofuranosidase from Geobacillus stearothermophilus T-6, a family 51 glycoside hydrolase. J Biol Chem 277:43667–43673

    Article  CAS  Google Scholar 

  • Spagna G, Romagnoli D, Angela M, Bianchi G, Pifferi PG (1998) A simple method for purifying glycosidases: α-l-arabinofuranosidase and β-d-glucopyranosidase from Aspergillus niger to increase the aroma of wine. Enzyme Microb Technol 22:298–304

    Article  CAS  Google Scholar 

  • Van Laere KM, Hartemink R, Bosveld M, Schols HA, Voragen AG (2000) Fermentation of plant cell wall derived polysaccharides and their corresponding oligosaccharides by intestinal bacteria. J Agric Food Chem 48:1644–1652

    Article  CAS  Google Scholar 

  • Ward OP, Moo-Young M (1989) Degradation of cell wall and related plant polysaccharides. CRC Crit Rev Biotechnol 8:237–274

    Article  CAS  Google Scholar 

  • Weinstein L, Albersheim P (1979) Structure of plant cell walls. IX. Purification and partial characterisation of a wall degrading endo-arabanase and an arabinosidase from Bacillus subtilis. Plant Physiol 63:425–432

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We are grateful to The Scientific and Research Council of Turkey (TUBITAK, Grant no. 104T286) and Karadeniz Technical University Research Foundation (grant no. 2007.111.04.10) for financial support.

Author information

Authors and Affiliations

  1. Department of Biology, Faculty of Arts and Sciences, Karadeniz Technical University, 61080, Trabzon, Turkey

    Sabriye Canakci, Murat Kacagan, Kadriye Inan & Ali Osman Belduz

  2. Fermentation Biotechnology Research Unit, National Center for Agricultural Utilization Research, USDA-ARS, 1815 N. University St., Peoria, IL, 61604, USA

    Badal C. Saha

Authors
  1. Sabriye Canakci
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Murat Kacagan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kadriye Inan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ali Osman Belduz
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Badal C. Saha
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Sabriye Canakci.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Canakci, S., Kacagan, M., Inan, K. et al. Cloning, purification, and characterization of a thermostable α-l-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari. Appl Microbiol Biotechnol 81, 61–68 (2008). https://doi.org/10.1007/s00253-008-1584-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-008-1584-1

Keywords

Search

Navigation