Cloning, expression, and characterization of a DNA ligase from a hyperthermophilic archaeon Thermococcus sp

Biotechnol Lett. 2006 Mar;28(6):401-7. doi: 10.1007/s10529-005-6070-6.

Abstract

Genomic analysis of a hyperthermophilic archaeon, Thermococcus sp. NA1, revealed an ORF of 1689 bases encoding 562 amino acids that showed a high similarity to DNA ligases from other hyperthermophilic archaea. The ligase, which was designated TNA1_lig (Thermococcus sp. NA1 ligase), was cloned and expressed in Escherichia coli. The recombinant TNA1_lig was purified by metal affinity chromatography. The optimum ligase activity of the recombinant TNA1_lig occurred at 80 degrees C and pH 7.5. The enzyme was activated by MgCl2 and ZnCl2 but was inhibited by MnCl2 and NiCl2. Additionally, the enzyme was activated by either ATP or NAD+.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Amino Acid Sequence
  • Bacteriological Techniques / methods
  • Gene Expression / genetics
  • Molecular Sequence Data
  • NAD / metabolism
  • Open Reading Frames / genetics
  • Thermococcus / enzymology*
  • Thermococcus / genetics

Substances

  • NAD
  • Adenosine Triphosphate