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Heterologous expression and characterization of Bacillus coagulans l-arabinose isomerase

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Abstract

Bacillus coagulans has been of great commercial interest over the past decade owing to its strong ability of producing optical pure l-lactic acid from both hexose and pentose sugars including l-arabinose with high yield, titer and productivity under thermophilic conditions. The l-arabinose isomerase (L-AI) from Bacillus coagulans was heterologously over-expressed in Escherichia coli. The open reading frame of the L-AI has 1,422 nucleotides encoding a protein with 474 amino acid residues. The recombinant L-AI was purified to homogeneity by one-step His-tag affinity chromatography. The molecular mass of the enzyme was estimated to be 56 kDa by SDS-PAGE. The enzyme was most active at 70°C and pH 7.0. The metal ion Mn2+ was shown to be the best activator for enzymatic activity and thermostability. The enzyme showed higher activity at acidic pH than at alkaline pH. The kinetic studies showed that the K m, V max and k cat/K m for the conversion of l-arabinose were 106 mM, 84 U/mg and 34.5 mM−1min−1, respectively. The equilibrium ratio of l-arabinose to l-ribulose was 78:22 under optimal conditions. l-ribulose (97 g/L) was obtained from 500 g/l of l-arabinose catalyzed by the enzyme (8.3 U/mL) under the optimal conditions within 1.5 h, giving at a substrate conversion of 19.4% and a production rate of 65 g L−1 h−1.

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References

  • 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 

  • Bucke C (1983) Practicality of industrial enzymes. Biochem Soc Trans 11:13–14

    CAS  Google Scholar 

  • Cheng L, Mu W, Zhang T, Jiang B (2010) An l-arabinose isomerase from Acidothermus cellulolytics ATCC 43068: cloning, expression, purification, and characterization. Appl Microbiol Biotechnol 86:1089–1097

    Article  CAS  Google Scholar 

  • Chouayekh H, Bejar W, Rhimi M, Jelleli K, Mseddi M, Bejar S (2007) Characterization of an l-arabinose isomerase from the Lactobacillus plantarum NC8 strain showing pronounced stability at acidic pH. FEMS Microbiol Lett 277:260–267

    Article  CAS  Google Scholar 

  • De Muynck C, Beauprez J, Soetaert W, Vandamme EJ (2006a) Boric acid as a mobile phase additive for high performance liquid chromatography separation of ribose, arabinose and ribulose. J Chromatogr A 1101:115–121

    Article  Google Scholar 

  • De Muynck C, Pereira C, Soetaert W, Vandamme E (2006b) Dehydrogenation of ribitol with Gluconobacter oxydans: production and stability of l-ribulose. J Biotechnol 125:408–415

    Article  Google Scholar 

  • Dische Z, Borenfreund E (1951) A new spectrophotometric method for the detection and determination of keto sugars and trioses. J Biol Chem 192:583–587

    CAS  Google Scholar 

  • Heath EC, Horecker BL, Smyrniotis PZ, Takagi Y (1958) Pentose fermentation by Lactobacillus plantarum. II. l-arabinose isomerase. J Biol Chem 231:1031–1037

    CAS  Google Scholar 

  • Helanto M, Kiviharju K, Leisola M, Nyyssola A (2007) Metabolic engineering of Lactobacillus plantarum for production of l-ribulose. Appl Environ Microbiol 73:7083–7091

    Article  CAS  Google Scholar 

  • Jorgensen F, Hansen OC, Stougaard P (2004) Enzymatic conversion of d-galactose to d-tagatose: heterologous expression and characterisation of a thermostable l-arabinose isomerase from Thermoanaerobacter mathranii. Appl Microbiol Biotechnol 64:816–822

    Article  CAS  Google Scholar 

  • Kim P (2004) Current studies on biological tagatose production using l-arabinose isomerase: a review and future perspective. Appl Microbiol Biotechnol 65:243–249

    CAS  Google Scholar 

  • Kim HJ, Oh DK (2005) Purification and characterization of an l-arabinose isomerase from an isolated strain of Geobacillus thermodenitrificans producing d-tagatose. J Biotechnol 120:162–173

    Article  CAS  Google Scholar 

  • Kim BC, Lee YH, Lee HS, Lee DW, Choe EA, Pyun YR (2002) Cloning, expression and characterization of l-arabinose isomerase from Thermotoga neapolitana: bioconversion of d-galactose to d-tagatose using the enzyme. FEMS Microbiol Lett 212:121–126

    CAS  Google Scholar 

  • Kim HJ, Kim JH, Oh HJ, Oh DK (2006) Characterization of a mutated Geobacillus stearothermophilus l-arabinose isomerase that increases the production rate of d-tagatose. J Appl Microbiol 101:213–221

    Article  CAS  Google Scholar 

  • Kim JH, Prabhu P, Jeya M, Tiwari MK, Moon HJ, Singh RK, Lee JK (2010) Characterization of an l-arabinose isomerase from Bacillus subtilis. Appl Microbiol Biotechnol 85:1839–1847

    Article  CAS  Google Scholar 

  • Kylma AK, Granstrom T, Leisola M (2004) Growth characteristics and oxidative capacity of Acetobacter aceti IFO 3281: implications for l-ribulose production. Appl Microbiol Biotechnol 63:584–591

    Article  CAS  Google Scholar 

  • Lee DW, Jang HJ, Choe EA, Kim BC, Lee SJ, Kim SB, Hong YH, Pyun YR (2004) Characterization of a thermostable l-arabinose (d-galactose) isomerase from the hyperthermophilic eubacterium Thermotoga maritima. Appl Environ Microbiol 70:1397–1404

    Article  CAS  Google Scholar 

  • Lee DW, Choe EA, Kim SB, Eom SH, Hong YH, Lee SJ, Lee HS, Lee DY, Pyun YR (2005a) Distinct metal dependence for catalytic and structural functions in the l-arabinose isomerases from the mesophilic Bacillus halodurans and the thermophilic Geobacillus stearothermophilus. Arch Biochem Biophys 434:333–343

    Article  CAS  Google Scholar 

  • Lee SJ, Lee DW, Choe EA, Hong YH, Kim SB, Kim BC, Pyun YR (2005b) Characterization of a thermoacidophilic l-arabinose isomerase from Alicyclobacillus acidocaldarius: role of Lys-269 in pH optimum. Appl Environ Microbiol 71:7888–7896

    Article  CAS  Google Scholar 

  • Liu SY, Wiegel J, Gherardini FC (1996) Purification and cloning of a thermostable xylose (glucose) isomerase with an acidic pH optimum from Thermoanaerobacterium strain JW/SL-YS 489. J Bacteriol 178:5938–5945

    CAS  Google Scholar 

  • Manjasetty BA, Chance MR (2006) Crystal structure of Escherichia coli l-arabinose isomerase (ECAI), the putative target of biological tagatose production. J Mol Biol 360:297–309

    Article  CAS  Google Scholar 

  • Mathe C, Gosselin G (2006) l-nucleoside enantiomers as antivirals drugs: a mini-review. Antiviral Res 71:276–281

    Article  CAS  Google Scholar 

  • Mizanur RM, Takata G, Izumori K (2001) Cloning and characterization of a novel gene encoding l-ribose isomerase from Acinetobacter sp. strain DL-28 in Escherichia coli. Biochim Biophys Acta 1521:141–145

    CAS  Google Scholar 

  • Oh DK (2007) Tagatose: properties, applications, and biotechnological processes. Appl Microbiol Biotechnol 76:1–8

    Article  CAS  Google Scholar 

  • Ou MS, Mohammed N, Ingram LO, Shanmugam KT (2009) Thermophilic Bacillus coagulans requires less cellulases for simultaneous saccharification and fermentation of cellulose to products than mesophilic microbial biocatalysts. Appl Biochem Biotechnol 155:379–385

    Article  CAS  Google Scholar 

  • Poonperm W, Takata G, Okada H, Morimoto K, Granstrom TB, Izumori K (2007) Cloning, sequencing, overexpression and characterization of l-rhamnose isomerase from Bacillus pallidus Y25 for rare sugar production. Appl Microbiol Biotechnol 76:1297–1307

    Article  CAS  Google Scholar 

  • Prabhu P, Tiwari MK, Jeya M, Gunasekaran P, Kim IW, Lee JK (2008) Cloning and characterization of a novel l-arabinose isomerase from Bacillus licheniformis. Appl Microbiol Biotechnol 81:283–290

    Article  CAS  Google Scholar 

  • Prabhu P, Jeya M, Lee JK (2010) Probing the molecular determinant for the catalytic efficiency of l-arabinose isomerase from Bacillus licheniformis. Appl Environ Microbiol 76:1653–1660

    Article  CAS  Google Scholar 

  • Qin J, Zhao B, Wang X, Wang L, Yu B, Ma Y, Ma C, Tang H, Sun J, Xu P (2009) Non-sterilized fermentative production of polymer-grade l-lactic acid by a newly isolated thermophilic strain Bacillus sp. 2–6. PLoS One 4:e4359

    Article  Google Scholar 

  • Rhimi M, Bejar S (2006) Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain. Biochim Biophys Acta 1760:191–199

    Article  CAS  Google Scholar 

  • Rhimi M, Juy M, Aghajari N, Haser R, Bejar S (2007) Probing the essential catalytic residues and substrate affinity in the thermoactive Bacillus stearothermophilus US100 l-arabinose isomerase by site-directed mutagenesis. J Bacteriol 189:3556–3563

    Article  CAS  Google Scholar 

  • Smith KW, Johnson SL (1976) Borate inhibition of yeast alcohol dehydrogenase. Biochemistry 15:560–565

    Article  CAS  Google Scholar 

  • Spagnuolo M, Crecchio C, Pizzigallo MD, Ruggiero P (1999) Fractionation of sugar beet pulp into pectin, cellulose, and arabinose by arabinases combined with ultrafiltration. Biotechnol Bioeng 64:685–691

    Article  CAS  Google Scholar 

  • Yeom SJ, Ji JH, Yoon RY, Oh DK (2008) l-Ribulose production from l-arabinose by an l-arabinose isomerase mutant from Geobacillus thermodenitrificans. Biotechnol Lett 30:1789–1793

    Article  CAS  Google Scholar 

  • Yeom SJ, Kim NH, Park CS, Oh DK (2009) l-ribose production from l-arabinose by using purified l-arabinose isomerase and mannose-6-phosphate isomerase from Geobacillus thermodenitrificans. Appl Environ Microbiol 75:6941–6943

    Article  CAS  Google Scholar 

  • Yoon SH, Kim P, Oh DK (2003) Properties of l-arabinose isomerase from Escherichia coli as biocatalyst for tagatose production. World J Microbiol Biotechnol 19:47–51

    Article  CAS  Google Scholar 

  • Zhang YW, Jeya M, Lee JK (2010) l-Ribulose production by an Escherichia coli harboring l-arabinose isomerase from Bacillus licheniformis. Appl Microbiol Biotechnol 87:1993–1999

    Article  CAS  Google Scholar 

  • Zhu W, Manjasetty BA, Chance MR (2007) Crystal structure of Mn2+-bound Escherichia coli l-arabinose Isomerase (ECAI) and Implications in protein catalytic mechanism and thermo-stability. J Young Investigators 17. http://www.jyi.org/research/re.php?id=1255. Accessed 7 Feb 2012

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Acknowledgments

This research was supported by the Science and Engineering Research Council (SERC) of the Agency for Science, Technology and Research (A*STAR) of Singapore (SERC grant no 0921590133). We are grateful to Dr. Keith Carpenter for critical reading of this manuscript.

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  1. Institute of Chemical and Engineering Sciences, Agency for Science, Technology and Research (A*STAR), 1 Pesek Road, Jurong Island, 627833, Singapore

    Xingding Zhou & Jin Chuan Wu

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  1. Xingding Zhou
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  2. Jin Chuan Wu
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Correspondence to Jin Chuan Wu.

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Zhou, X., Wu, J.C. Heterologous expression and characterization of Bacillus coagulans l-arabinose isomerase. World J Microbiol Biotechnol 28, 2205–2212 (2012). https://doi.org/10.1007/s11274-012-1026-1

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  • DOI: https://doi.org/10.1007/s11274-012-1026-1

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