Résumé
A thermostable DNA ligase gene was identified, and the biochemical and enzymatic properties of the ligase were characterized from JP2 strain which was enriched from geothermally active sites in Papua New Guinea. The nucleotide and amino acid sequences showed much high identities compared with that of archeabacterium species Sulfolobus solfataricus and Sulfolobus shibatae,especially in the six conserved motif sequences, which are known to be closely related to the key function of ligase. Recombinant JP2 ligase showed high activity in nick-joining reaction. It was the most active when Mn2+ present as divalent metal cofactor rather than Mg2+ and Ca2+ etc.. Assay of thermostability over a range of temperatures showed that at 50～80℃ the enzyme displayed relative high activity. Further thermostability experiment indicated that the activity of JP2 ligase could last for a long time at 80℃ and 85℃,however, at 90℃ and 95℃, it became unstable quickly. An investigation on the acquired thermotolerance of recombinant JP2 ligase was done by applying a chaperonin known as TF55 in thermophile on JP2 ligase reaction. Result showed that TF55 could not help in improving thermostability of ligase at 85℃. The possible reason might be that at 85℃ in vitro, the chaperonin itself was denatured.