Extremophiles

, Volume 5, Issue 2, pp 119–126

Cloning, expression, and characterization of a chitinase gene from the Antarctic psychrotolerant bacterium Vibrio sp. strain Fi:7

Authors

  • Anne Bendt
    • Institute of Marine Biotechnology, W.-Rathenau Str. 49a, D-17489 Greifswald, Germany
  • Heike Hüller
    • Institute of Marine Biotechnology, W.-Rathenau Str. 49a, D-17489 Greifswald, Germany
  • Ulrike Kammel
    • Institute of Marine Biotechnology, W.-Rathenau Str. 49a, D-17489 Greifswald, Germany
  • Elisabeth Helmke
    • Alfred Wegener Institute for Polar and Marine Research, Bremerhaven, Germany
  • Thomas Schweder
    • Institute of Marine Biotechnology, W.-Rathenau Str. 49a, D-17489 Greifswald, Germany
Original paper

DOI: 10.1007/s007920100179

Cite this article as:
Bendt, A., Hüller, H., Kammel, U. et al. Extremophiles (2001) 5: 119. doi:10.1007/s007920100179

Abstract.

A marine psychrotolerant bacterium from the Antarctic Ocean showing high chitinolytic activity on chitin agar at 5°C was isolated. The sequencing of the 16S rRNA indicates taxonomic affiliation of the isolate Fi:7 to the genus Vibrio. By chitinase activity screening of a genomic DNA library of Vibrio sp. strain Fi:7 in Escherichia coli, three chitinolytic clones could be isolated. Sequencing revealed, for two of these clones, the same open reading frame of 2,189 nt corresponding to a protein of 79.4 kDa. The deduced amino acid sequence of the open reading frame showed homology of 82% to the chitinase ChiA from Vibrio harveyi. The chitinase of isolate Fi:7 contains a signal peptide of 26 amino acids. Sequence alignment with known chitinases showed that the enzyme has a chitin-binding domain and a catalytic domain typical of other bacterial chitinases. The chitinase ChiA of isolate Fi:7 was overexpressed in E. coli BL21(DE3) and purified by anion-exchange and hydrophobic interaction chromatography. Maximal enzymatic activity was observed at a temperature of 35°C and pH 8. Activity of the chitinase at 5°C was 40% of that observed at 35°C. Among the main cations contained in seawater, i.e., Na+, K+, Ca2+, and Mg2+, the enzymatic activity of ChiA could be enhanced twofold by the addition of Ca2+.

Chitinase Cold adapted Marine bacterium Psychrotolerant Vibrio sp.
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© Springer-Verlag 2001