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Cloning, expression, and characterization of a glycoside hydrolase family 50 β-agarase from a marine Agarivorans isolate

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Abstract

The gene for a thermostable β-agarase from Agarivorans sp. JA-1 was cloned and sequenced. It comprised an open reading frame of 2,988 base pairs, which encode a protein of 109,450 daltons consisting of 995 amino acid residues. A comparison of the entire sequence showed that the enzyme has 98.8% sequence similarities to β-agarase from Vibrio sp. JT1070, indicating that it belongs to the family glycoside hydrolase (GH)-50. The gene corresponding to a mature protein of 976 amino acids was inserted and expressed in Escherichia coli. The recombinant β-agarase was purified to homogeneity. It had maximal activity at 40°C and pH 8.0 in the presence of 1 mM NaCl and 1 mM CaCl2. The enzyme hydrolyzed agarose as well as neoagarohexaose and neoagarotetraose to yield neoagarobiose as the main product. Thus, the enzyme would be useful for the industrial production of neoagarobiose.

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References

  • Allouch J, Jam M, Helbert W, Barbeyron T, Kloareg B, Henrissat B, Czjzek M (2003) The three-dimensional structures of two β-agarases. J Biol Chem 278:47171–47180

    Article  PubMed  CAS  Google Scholar 

  • Aoki T, Araki T, Kitamikado M (1990) Purification and characterization of a novel β-agarase from Vibrio sp. AP-2. Eur J Biochem 187:461–465

    Article  PubMed  CAS  Google Scholar 

  • Araki C (1959) Seaweed polysaccharides. In: Wolfrom ML (ed) Carbohydrate chemistry of substances of biological interest. Pergamon Press, London pp 15–30

    Google Scholar 

  • Belas R, Bartlett D, Michael S (1988) Cloning and gene replacement mutagenesis of a Pseudomonas atlantica agarase gene. Appl Environ Microbiol 54:30–37

    PubMed  CAS  Google Scholar 

  • Duckworth M, Yaphe W (1970) Thin-layer chromatographic analysis of enzymic hydrolysate of agar. J Chromatogr 49:482–487

    Article  PubMed  CAS  Google Scholar 

  • Duckworth M, Yaphe W (1971) Structure of agar. I. Fractionation of a complex mixture of polysaccharides. Carbohydr Res 16:189–197

    Article  CAS  Google Scholar 

  • Groleau D, Yaphe W (1977) Enzymatic hydrolysis of agar: purification and characterization of β-neoagarotetraose hydrolase from Peudomonas atlantica. Can J Microbiol 23:672–679

    Article  PubMed  CAS  Google Scholar 

  • Ha JC, Kim GT, Kim SK, Oh TK, Yu JH, Kong IS (1997) Beta-Agarase from Pseudomonas sp. W7: purification of the recombinant enzyme from Escherichia coli and the effects of salt on its activity. Biotechnol Appl Biochem 26:1–6

    PubMed  CAS  Google Scholar 

  • Kim BJ, Kim HJ, Ha SD, Hwang SH, Byun DS, Lee TH, Kong JY (1999) Purification and characterization of β-agarase from marine bacterium Bacillus cereus ASK202. Biotechnol Lett 21:1011–1105

    Article  CAS  Google Scholar 

  • Kobayashi R, Takisada M, Suzuki T, Kirimura K, Usami S (1997) Neoagarobiose as a novel moisturizer with whitening effect. Biosci Biotechnol Biochem 61:62–63

    Google Scholar 

  • Kohno T, Kitagawa H, Hiraga T (1990) Production of hetero-oligosaccharides. In: Gijutsu Kenkyu Kukami (eds) Shokuhin sangyo bioreactor system, Jissen bioreactor, Shokuhin Kagaku Shimbunsa, Tokyo, Japan, pp 87–105

  • Kono T, Hidaka H (1989) Properties and production of neoagarooligosaccharide. Nippon Nogeikagaku Kaishi 63:1126–1129

    CAS  Google Scholar 

  • Kurahashi M, Yokota A (2004) Agarivorans albus gen. nov., sp. nov., a gamma-proteobacterium isolated from marine animals. Int J Syst Evol Microbiol 54:693–697

    Article  PubMed  CAS  Google Scholar 

  • Ohta Y, Nogi Y, Miyazaki M, Li Z, Hatada Y, Ito S, Horikoshi K (2004) Enzymatic properties and nucleotide and amino acid sequences of a thermostable beta-agarase from the novel marine isolate, JAMB-A94. Biosci Biotechnol Biochem 68:1073–1081

    Article  PubMed  CAS  Google Scholar 

  • Sambrook E, Fritsch F, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Labarotory, Cold Spring Harbor, New York

    Google Scholar 

  • Schroeder DC, Jaffer MA, Coyne VE (2003) Investigation of the role of a beta(1-4) agarase produced by Pseudoalteromonas gracilis B9 in eliciting disease symptoms in the red alga Gracilaria gracilis. Microbiology 149:2919–2929

    Article  PubMed  CAS  Google Scholar 

  • Somogyi M (1952) Notes on sugar determination. J Biol Chem 195:19–23

    CAS  Google Scholar 

  • Sugano Y, Matsumoto T, Kodama H, Noma M (1993) Cloning and sequencing of agaA, a unique agarase 0107 gene from a marine bacterium, Vibrio sp. strain JT0107. Appl Environ Microbiol 59:3750–3756

    PubMed  CAS  Google Scholar 

  • Yoshizawa Y, Ametani A, Tsunehiro J, Nomura K, Itoh M, Fukui F, Kaminogawa S (1995) Macrophage stimulation activity of the polysaccharide fraction from a marine alga (Porphyra yezoensis): structure–function relationships and improved solubility. Biosci Biotechnol Biochem 59:1933–1937

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgement

This work was supported by the Marine and Extreme Genome Research Center Program, Ministry of Maritime Affairs & Fisheries, Republic of Korea.

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Authors and Affiliations

  1. Department of Pharmaceutical Engineering, College of Medical Life Sciences, Silla University, Busan, 617-736, Korea

    Dong-Geun Lee, Nam Young Kim, Eo-Jin Lee, Min Kyung Jang, Young Gyun Shin, Gwang-Seok Park, Tae-Min Kim, Jae-Hwa Lee & Sang-Hyeon Lee

  2. Research and University-Industry Cooperation, Pusan National University, Busan, 609-735, Korea

    Geun-Tae Park

  3. Microbiology Laboratory, Korea Ocean Research and Development Institute, PO box 29, Ansan, 425-600, Korea

    Jung-Hyun Lee & Sang-Jin Kim

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  1. Dong-Geun Lee
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  2. Geun-Tae Park
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  3. Nam Young Kim
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Correspondence to Sang-Hyeon Lee.

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Lee, DG., Park, GT., Kim, N.Y. et al. Cloning, expression, and characterization of a glycoside hydrolase family 50 β-agarase from a marine Agarivorans isolate. Biotechnol Lett 28, 1925–1932 (2006). https://doi.org/10.1007/s10529-006-9171-y

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  • DOI: https://doi.org/10.1007/s10529-006-9171-y

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