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Characterization of an L-arabinose isomerase from Bacillus subtilis

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Abstract

An isolated gene from Bacillus subtilis str. 168 encoding a putative isomerase was proposed as an L-arabinose isomerase (L-AI), cloned into Escherichia coli, and its nucleotide sequence was determined. DNA sequence analysis revealed an open reading frame of 1,491 bp, capable of encoding a polypeptide of 496 amino acid residues. The gene was overexpressed in E. coli and the protein was purified using nickel-nitrilotriacetic acid chromatography. The purified enzyme showed the highest catalytic efficiency ever reported, with a k cat of 14,504 min−1 and a k cat/K m of 121 min−1 mM−1 for L-arabinose. A homology model of B. subtilis L-AI was constructed based on the X-ray crystal structure of E. coli L-AI. Molecular dynamics simulation studies of the enzyme with the natural substrate, L-arabinose, and an analogue, D-galactose, shed light on the unique substrate specificity displayed by B. subtilis L-AI only towards L-arabinose. Although L-AIs have been characterized from several other sources, B. subtilis L-AI is distinguished from other L-AIs by its high substrate specificity and catalytic efficiency for L-arabinose.

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References

  • Akagi M, Omae D, Tamura Y, Ueda T, Kumashiro T, Urata H (2002) A practical synthesis of L-ribose. Chem Pharm Bull (Tokyo) 50:866–868

    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 

  • Brooks B, Bruccoleri RE, Olafson BD, States DJ, Swaminathan S, Karplus M (1983) CHARMm: a program for macromolecular energy, minimization, and dynamics calculations. J Comput Chem 4:187–217

    Article  CAS  Google Scholar 

  • Cho B, Kim JH, Jeon HB, Kim KS (2005) A new efficient and practical synthesis of 2-deoxy-L-ribose. Tetrahedron 61:4341–4346

    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 

  • 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 

  • Izumori K (2002) Bioproduction strategies for rare hexose sugars. Naturwissenschaften 89:120–124

    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 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 

  • Kunst F, Ogasawara N, Moszer I, Albertini AM, Alloni G, Azevedo V, Bertero MG, Bessieres P, Bolotin A, Borchert S et al (1997) The complete genome sequence of the gram-positive bacterium Bacillus subtilis. Nature 390:249–256

    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 DW, Hong YH, Choe EA, Lee SJ, Kim SB, Lee HS, Oh JW, Shin HH, Pyun YR (2005b) A thermodynamic study of mesophilic, thermophilic, and hyperthermophilic L-arabinose isomerases: the effects of divalent metal ions on protein stability at elevated temperatures. FEBS Lett 579:1261–1266

    Article  CAS  Google Scholar 

  • Lee SJ, Lee DW, Choe EA, Hong YH, Kim SB, Kim BC, Pyun YR (2005c) 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 

  • 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 

  • Patrick JW, Lee N (1968) Purification and properties of an L-arabinose isomerase from Escherichia coli. J Biol Chem 243:4312–4318

    CAS  Google Scholar 

  • Prabhu P, Tiwari M, 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 

  • 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

    CAS  Google Scholar 

  • Roh HJ, Kim P, Park YC, Choi JH (2000) Bioconversion of D-galactose into D-tagatose by expression of L-arabinose isomerase. Biotechnol Appl Biochem 31(Pt 1):1–4

    Article  CAS  Google Scholar 

  • Seemann JE, Schulz GE (1997) Structure and mechanism of L-fucose isomerase from Escherichia coli. J Mol Biol 273:256–268

    Article  CAS  Google Scholar 

  • Wu G, Robertson DH, Brooks CL 3rd, Vieth M (2003) Detailed analysis of grid-based molecular docking: a case study of CDOCKER-A CHARMm-based MD docking algorithm. J Comput Chem 24:1549–1562

    Article  CAS  Google Scholar 

  • Yoon S, 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 

  • Yun M, Moon HR, Kim HO, Choi WJ, Kim YC, Park CS, Jeong LS (2005) A highly efficient synthesis of unnatural l-sugars from d-ribose. Tetrahedron Lett 46:5903–5905

    Article  CAS  Google Scholar 

  • Zakaria A (2001) Production of natural and rare pentoses using microorganisms and their enzymes. Electron J Biotechnol 4:103–111

    Google Scholar 

Download references

Acknowledgment

This work was supported by the 21C Frontier Microbial Genomics and Applications Center Program, Ministry of Education, Science & Technology, Republic of Korea. It was also supported by a grant (code 2008A0080126) from Agricultural Research and Development Promotion Center and a grant (code 20070301034024) from the Biogreen 21 Program, Rural Development Administration, Republic of Korea.

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Authors and Affiliations

  1. BioNgene Co. Ltd., 10-1 Myungryun-Dong, Jongro-Gu, Seoul, 110521, Korea

    Jin-Ha Kim

  2. Department of Bioscience and Biotechnology, Konkuk University, 1 Hwayang-Dong, Gwangjin-Gu, Seoul, 143701, Korea

    Ponnandy Prabhu & Hee-Jung Moon

  3. Department of Chemical Engineering, Konkuk University, 1 Hwayang-Dong, Gwangjin-Gu, Seoul, 143701, Korea

    Marimuthu Jeya, Manish Kumar Tiwari, Raushan Kumar Singh & Jung-Kul Lee

  4. Institute of Biomedical Science and Technology, Konkuk University, 1 Hwayang-Dong, Gwangjin-Gu, Seoul, 143701, Korea

    Jung-Kul Lee

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  1. Jin-Ha Kim
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  2. Ponnandy Prabhu
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Correspondence to Jung-Kul Lee.

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Kim, JH., Prabhu, P., Jeya, M. et al. Characterization of an L-arabinose isomerase from Bacillus subtilis . Appl Microbiol Biotechnol 85, 1839–1847 (2010). https://doi.org/10.1007/s00253-009-2210-6

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  • DOI: https://doi.org/10.1007/s00253-009-2210-6

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