Extremophiles

, Volume 5, Issue 1, pp 35–44

Cloning of two pectate lyase genes from the marine Antarctic bacterium Pseudoalteromonas haloplanktis strain ANT/505 and characterization of the enzymes

  • Le Van Truong
  • Ho Tuyen
  • Elisabeth Helmke
  • Le Tran Binh
  • Thomas Schweder
Original paper

DOI: 10.1007/s007920000170

Cite this article as:
Van Truong, L., Tuyen, H., Helmke, E. et al. Extremophiles (2001) 5: 35. doi:10.1007/s007920000170

Abstract

A marine Antarctic psychrotolerant bacterium (strain ANT/505), isolated from sea ice-covered surface water from the Southern Ocean, showed pectinolytic activity on citrus pectin agar. The sequencing of the 16S rRNA of isolate ANT/505 indicates a taxonomic affiliation to Pseudoalteromonas haloplanktis. The supernatant of this strain showed three different pectinolytic activities after growth on citrus pectin. By activity screening of a genomic DNA library of isolate ANT/505 in Escherichia coli, two different pectinolytic clones could be isolated. Subcloning and sequencing revealed two open reading frames (ORF) of 1,671 and 1,968 nt, corresponding to proteins of 68 and 75 kDa, respectively. The deduced amino acid sequence of the two ORFs showed homology to pectate lyases from Erwinia chrysanthemi and Aspergillus nidulans. The pectate lyases contain signal peptides of 17 and 26 amino acids that were correctly processed after overexpression in E. coli BL21. Both enzymes were purified by anionic exchange chromatography. Maximal enzymatic activities for both pectate lyases were observed at 30°C and a pH range of 9 to 10. The Km values of both lyases for pectate and citrus pectin were 1 g l–1 and 5 g l–1, respectively. Calcium was required for activity on pectic substrates, whereas the addition of 1 mM ethylenediaminetetraacetic acid (EDTA) resulted in complete inhibition of the enzymes. These two enzymes represent the first pectate lyases isolated and characterized from a cold-adapted marine bacterium.

Pectate lyase Pectin Psychrophilic Marine Pseudoalteromonas haloplanktis 

Copyright information

© Springer-Verlag 2001

Authors and Affiliations

  • Le Van Truong
    • 1
  • Ho Tuyen
    • 1
  • Elisabeth Helmke
    • 2
  • Le Tran Binh
    • 1
  • Thomas Schweder
    • 3
  1. 1.Institute of Biotechnology, National Center of Natural Science and Technology, TuLiem, HanoiVietnam
  2. 2.Alfred Wegener Institute for Polar and Marine Research, BremerhavenGermany
  3. 3.Institute of Marine Biotechnology, W.-Rathenau Str. 49a, D-17489 GreifswaldGermany

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