Comparative analyses of the two proliferating cell nuclear antigens from the hyperthermophilic archaeon, Thermococcus kodakarensis.
The DNA sliding clamp is a multifunctional protein involved in cellular DNA transactions. In Archaea and Eukaryota, proliferating cell nuclear antigen (PCNA) is the sliding clamp. The ring-shaped PCNA encircles double-stranded DNA within its central hole and tethers other proteins on DNA. The majority of Crenarchaeota, a subdomain of Archaea, have multiple PCNA homologues, and they are capable of forming heterotrimeric rings for their functions. In contrast, most organisms in Euryarchaeota, the other major subdomain, have a single PCNA forming a homotrimeric ring structure. Among the Euryarchaeota whose genome is sequenced, Thermococcus kodakarensis is the only species with two genes encoding PCNA homologues on its genome. We cloned the two genes from the T. kodakarensis genome, and the gene products, PCNA1 and PCNA2, were characterized. PCNA1 stimulated the DNA synthesis reactions of the two DNA polymerases, PolB and PolD, from T. kodakarensis in vitro. PCNA2, however, only had an effect on PolB. We were able to disrupt the gene for PCNA2, whereas gene disruption for PCNA1 was not possible, suggesting that PCNA1 is essential for DNA replication. The sensitivities of the Δpcna2 mutant strain to ultraviolet irradiation (UV), methyl methanesulfonate (MMS) and mitomycin C (MMC) were indistinguishable from those of the wild-type strain.
Department of Bioscience & Biotechnology, Faculty of Agriculture and Graduate School of Bioresource & Bioenvironmental Sciences, Kyushu University, Fukuoka, 812-8581, Japan.
Yumani
Y
Kuba
Sonoko
S
Ishino
Takeshi
T
Yamagami
Masahiro
M
Tokuhara
Tamotsu
T
Kanai
Ryosuke
R
Fujikane
Hiromi
H
Daiyasu
Haruyuki
H
Atomi
Yoshizumi
Y
Ishino
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004249:DNA Damage; D004258:DNA Polymerase III / Q000737:chemistry; D019951:DNA Polymerase beta / Q000737:chemistry; D004260:DNA Repair; D004261:DNA Replication; D019641:DNA, Archaeal / Q000737:chemistry / Q000378:metabolism; D055786:Gene Knockout Techniques; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D021122:Protein Subunits / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D051818:Replication Protein C / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D019712:Thermococcus / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*
D019843:Archaeal Proteins; D019641:DNA, Archaeal; D018809:Proliferating Cell Nuclear Antigen; D021122:Protein Subunits; D004258:DNA Polymerase III; D019951:DNA Polymerase beta; D000251:Adenosine Triphosphatases; D051818:Replication Protein C