Advertisement

Current Microbiology

, Volume 50, Issue 4, pp 212–216 | Cite as

Purification and Characterization of a Novel α-Agarase from a Thalassomonas sp.

  • Yukari Ohta
  • Yuji HatadaEmail author
  • Masayuki Miyazaki
  • Yuichi Nogi
  • Susumu Ito
  • Koki Horikoshi
Article

Abstract

An agar-degrading Thalassomonas bacterium, strain JAMB-A33, was isolated from the sediment off Noma Point, Japan, at a depth of 230 m. A novel α-agarase from the isolate was purified to homogeneity from cultures containing agar as a carbon source. The molecular mass of the purified enzyme, designated as agaraseA33, was 85 kDa on both SDS-PAGE and gel-filtration chromatography, suggesting that it is a monomer. The optimal pH and temperature for activity were about 8.5 and 45°C, respectively. The enzyme had a specific activity of 40.7 U/mg protein. The pattern of agarose hydrolysis showed that the enzyme is an endo-type α-agarase, and the final main product was agarotetraose. The enzyme degraded not only agarose but also agarohexaose, neoagarohexaose, and porphyran.

Keywords

Standard Assay Condition Agarase Activity Lysyl Endopeptidase BioWhittaker Molecular Application Sulfated Agarose 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

We are grateful to Dr. Y. Sakano of Tokyo University of Agriculture and Technology for stimulating discussions.

Literature Cited

  1. 1.
    Allouch, J, Jam, M, Helbert, W, Barbeyron, T, Kloareg, B, Henrissat, B,  et al. 2003The three-dimensional structures of two β-agarasesJ Biol Chem2784717147180CrossRefPubMedGoogle Scholar
  2. 2.
    Duckworth, M, Yaphe, W 1971Structure of agar. I. Fractionation of a complex mixture of polysaccharidesCarbohydr Res1189197CrossRefGoogle Scholar
  3. 3.
    Ha, JC, Kim, GT, Kim, SK, Oh, TK, Yu, JH, Kong, IS 1997β-agarase from Pseudomonas sp. W7: Purification of the recombinant enzyme from Escherichia coli and the effects of salt on its activityBiotechnol Appl Biochem2616PubMedGoogle Scholar
  4. 4.
    Hosoda, A, Sakai, M, Kanazawa, S 2003Isolation and characterization of agar-degrading Paenibacillus spp. associated with the rhizosphere of spinachBiosci Biotechnol Biochem6710481055CrossRefPubMedGoogle Scholar
  5. 5.
    Kato, I 2000Antioxidative and antitumorigenic properties of agaro-oligosaccharideBio Industry171319Google Scholar
  6. 6.
    Kendall, K, Cullum, J 1984Cloning and expression of an extracellular-agarase from Streptomyces coelicolor A3(2) in Streptomyces lividans 66Gene29315321CrossRefPubMedGoogle Scholar
  7. 7.
    Kim, BJ, Kim, HJ, Ha, SH, Hwang, SH, Byun, DS, Lee, TH,  et al. 1999Purification and characterization of β-agarase from marine bacterium Bacillus cereus ASK202Biotechnol Lett2110111015CrossRefGoogle Scholar
  8. 8.
    Kim, YH, Kim, YS, Lee, JR, Lee, EK, Seu, JH 1993Enzymatic characteristics of an extracellular agarase of Cytophaga sp. KY-1 and molecular cloning of the agarase geneJ Microbiol Biotechnol33138Google Scholar
  9. 9.
    Laemmli, UK 1970Cleavage of structural proteins during the assembly of the head of bacteriophage T4Nature227680685PubMedGoogle Scholar
  10. 10.
    Morrice, LM, McLean, MW, Long, WF, Williamson, FB 1983β-agarases I and II from Pseudomonas atlantica. Substrate specificitiesEur J Biochem137149154CrossRefPubMedGoogle Scholar
  11. 11.
    Naganuma, T, Coury, DA, Poline-Fuller, M, Gibor, A, Horikoshi, K 1993Characterization of agarolytic Microscilla isolates and their extracellular agarasesSyst Appl Microbiol16183190Google Scholar
  12. 12.
    Ohta, Y, Hatada, Y, Nogi, Y, Li, Z, Zhang, HM, Ito, S,  et al. 2004aThermostable β-agarase from a deep-sea Microbulbifer isolateJ Appl Glycosci51203210Google Scholar
  13. 13.
    Ohta, Y, Hatada, Y, Nogi, Y, Miyazaki, M, Li, Z, Akita, M,  et al. 2004bEnzymatic properties and nucleotide and amino acid sequences of a thermostable β-agarase from a novel species of deep-sea MicrobulbiferAppl Microbiol Biotechnol64505514CrossRefGoogle Scholar
  14. 14.
    Potin, P, Richard, C, Rochas, C, Kloareg, B 1993Purification and characterization of the α-agarase from Alteromonas agarlyticus (Cataldi) combnov., strain GJ1B. Eur J Biochem214599607Google Scholar
  15. 15.
    Rochas, C, Potin, P, Kloareg, B 1994NMR spectroscopic investigation of agarose oligomers produced by an α-agaraseCarbohydr Res2536977CrossRefPubMedGoogle Scholar
  16. 16.
    Schroeder, DC, Jaffer, MA, Coyne, VE 2003Investigation of the role of a β (1–4) agarase produced by Pseudoalteromonas gracilis B9 in eliciting disease symptoms in the red alga Gracilaria gracilisMicrobiology14929192929CrossRefPubMedGoogle Scholar
  17. 17.
    Sugano, Y, Kodama, H, Terada, I, Yamazaki, Y, Noma, M 1994Purification and characterization of a novel enzyme, α-neoagarooligosaccharide hydrolase (α-NAOS hydrolase), from a marine bacterium, Vibrio sp. strain JT0107J Bacteriol17668126818PubMedGoogle Scholar
  18. 18.
    Sugano, Y, Terada, I, Noma, M, Matsumoto, T 1993Purification and characterization of a new agarase from a marine bacterium, Vibrio sp. strain JT0107Appl Environ Microbiol591549 1554Google Scholar
  19. 19.
    Takahashi, K, Hirano, Y, Araki, S, Hattori, M 2000Emulsifying ability of porphyran prepared from dried nori, Porphyra yezoensis, a red algaJ Agric Food Chem4827212725CrossRefPubMedGoogle Scholar
  20. 20.
    Weisburg, WG, Barns, SM, Pelletier, DA, Lane, DJ 199116S ribosomal DNA amplification for phylogenetic studyJ Bacteriol173697703PubMedGoogle Scholar
  21. 21.
    Winter, WP, Yodh, J 1983Interaction of human hemoglobin and its variants with agarScience221175178PubMedGoogle Scholar
  22. 22.
    Yoshizawa, Y, Ametani, A, Tsunehiro, J, Nomura, K, Itoh, M, Fukui, F,  et al. 1995Macrophage stimulation activity of the polysaccharide fraction from a marine alga (Porphyra yezoensis): structure–function relationships and improved solubilityBiosci Biotechnol Biochem5919331937PubMedGoogle Scholar
  23. 23.
    Zhang, Q, Li, N, Liu, X, Zhao, Z, Li, Z, Xu, Z 2004The structure of a sulfated galactan from Porphyra haitanensis and its in vivo antioxidant activityCarbohydr Res339105111CrossRefPubMedGoogle Scholar
  24. 24.
    Zhong, Z, Toukdarian, A, Helinski, D, Knauf, V, Sykes, S, Wilkinson, JE,  et al. 2001Sequence analysis of a 101-kilobase plasmid required for agar degradation by a Microscilla isolateAppl Environ Microbiol6757715779Google Scholar

Copyright information

© Springer Science+Business Media, Inc. 2005

Authors and Affiliations

  • Yukari Ohta
    • 1
  • Yuji Hatada
    • 1
    Email author
  • Masayuki Miyazaki
    • 1
  • Yuichi Nogi
    • 1
  • Susumu Ito
    • 1
  • Koki Horikoshi
    • 1
  1. 1.Japan Agency for Marine-Earth Science and Technology (JAMSTEC)Yokosuka Japan

Personalised recommendations