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Purification and characterization of a novel β-agarase, AgaA34, from Agarivorans albus YKW-34

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Abstract

An extracellular β-agarase (AgaA34) was purified from a newly isolated marine bacterium, Agarivorans albus YKW-34 from the gut of a turban shell. AgaA34 was purified to homogeneity by ion exchange and gel filtration chromatographies with a recovery of 30% and a fold of ten. AgaA34 was composed of a single polypeptide chain with the molecular mass of 50 kDa. N-terminal amino acid sequencing revealed a sequence of ASLVTSFEEA, which exhibited a high similarity (90%) with those of agarases from glycoside hydrolase family 50. The pH and temperature optima of AgaA34 were pH 8.0 and 40°C, respectively. It was stable over pH 6.0–11.0 and at temperature up to 50°C. Hydrolysis of agarose by AgaA34 produced neoagarobiose (75 mol%) and neoagarotetraose (25 mol%), whose structures were identified by matrix-assisted laser desorption ionization time-of-flight mass spectroscopy and 13C NMR. AgaA34 cleaved both neoagarohexaose and neoagarotetraose into neoagarobiose. The k cat/K m values for hydrolysis agarose and neoagarotetraose were 4.04 × 103 and 8.1 × 102 s−1 M−1, respectively. AgaA34 was resistant to denaturing reagents (sodium dodecyl sulfate and urea). Metal ions were not required for its activity, while reducing reagents (β-Me and dithiothreitol, DTT) increased its activity by 30%.

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Acknowledgments

This work was supported by the grant of No. RTI05-01-02 from the Regional Technology Innovation Program of the Ministry of Commerce, Industry and Energy (MOCIE), and Kangnung National University, Korea. Xiao Ting Fu was a recipient of a graduate fellowship provided by Brain Korea (BK21) program sponsored by the ministry of Education and Human Resource Development, Korea. We would like to thank Professor Il Shik Shin (Faculty of Marine Bioscience & Technology, Kangnung National University, Korea) for giving us the strain of Agarivorans albus YKW-34.

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Correspondence to Sang Moo Kim.

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Fu, X.T., Lin, H. & Kim, S.M. Purification and characterization of a novel β-agarase, AgaA34, from Agarivorans albus YKW-34. Appl Microbiol Biotechnol 78, 265–273 (2008). https://doi.org/10.1007/s00253-007-1303-3

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Keywords

  • β-Agarase
  • Agarivorans albus
  • Purification
  • Characterization
  • Neoagarobiose