Gene cloning and function analysis of replication factor C from Thermococcus kodakaraensis KOD1.
Replication factor C (RFC) catalyzes the assembly of circular proliferating cell nuclear antigen (PCNA) clamps around primed DNA, enabling processive synthesis by DNA polymerase. The RFC-like genes, arranged in tandem in the Thermococcus kodakaraensis KOD1 genome, were cloned individually and co-expressed in Escherichia coli cells. T. kodakaraensis KOD1 RFC homologue (Tk-RFC) consists of the small subunit (Tk-RFCS: MW=37.2 kDa) and the large subunit (Tk-RFCL: MW=57.2 kDa). The DNA elongation rate of the family B DNA polymerase from T. kodakaraensis KOD1 (KOD DNA polymerase), which has the highest elongation rate in all thermostable DNA polymerases, was increased about 1.7 times, when T. kodakaraensis KOD1 PCNA (Tk-PCNA) and the Tk-RFC at the equal molar ratio of KOD DNA polymerase were reacted with primed DNA.
Kitabayashi
Masao
M
Tsuruga Institute of Biotechnology, Toyobo Co., Ltd., Tsuruga, Fukui, Japan. masao_kitabayashi@bio.toyobo.co.jp
Nishiya
Yoshiaki
Y
Esaka
Muneharu
M
Itakura
Mitsuo
M
Imanaka
Tadayuki
T
D000595:Amino Acid Sequence; D001105:Archaea / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence; D017931:DNA Primers; D004261:DNA Replication / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D016680:Genome, Bacterial; D008969:Molecular Sequence Data; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics; D051818:Replication Protein C; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D019712:Thermococcus / Q000235:genetics*
D017931:DNA Primers; D004268:DNA-Binding Proteins; D018809:Proliferating Cell Nuclear Antigen; D051818:Replication Protein C