Cloning, expression, and characterization of DNA polymerase I from the hyperthermophilic archaea Thermococcus fumicolans.
Extremophiles
; 4(4): 215-25, 2000 Aug.
Article
en En
| MEDLINE
| ID: mdl-10972190
ABSTRACT
The DNA polymerase I gene of a newly described deep-sea hydrothermal vent Archaea species, Thermococcus fumicolans, from IFREMERS's collection of hyperthermophiles has been cloned in Escherichia coli. As in Thermococcus litoralis, the gene is split by two intervening sequences (IVS) encoding inteins inserted in sites A and C of family B DNA polymerases. The entire DNA polymerase gene, containing both inteins, was expressed at 30 degrees C in E. coli strain BL21(DE3)pLysS using the pARHS2 expression vector. The native polypeptide precursor of 170kDa was obtained, and intein splicing as well as ligation of the three exteins was observed in vitro after heat exposure. The recombinant enzyme was purified and some of its activities were characterized polymerization, thermostability, exonuclease activities, and fidelity.
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Colección:
01-internacional
Asunto principal:
Thermococcus
/
ADN Polimerasa I
Idioma:
En
Revista:
Extremophiles
Asunto de la revista:
BIOLOGIA
Año:
2000
Tipo del documento:
Article
País de afiliación:
Francia