Molecular analyses of an unusual translesion DNA polymerase from Methanosarcina acetivorans C2A.
The domain Archaea is composed of several subdomains, and prominent among them are the Crenarchaeota and the Euryarchaeota. Biochemically characterized archaeal family Y DNA polymerases (Pols) or DinB homologs, to date, are all from crenarchaeal organisms, especially the genus Sulfolobus. Here, we demonstrate that archaeal family Y Pols fall into five clusters based on phylogenetic analysis. MacDinB-1, the homolog from the euryarchaeon Methanosarcina acetivorans that is characterized in this study, belongs to cluster II. Therefore, MacDinB-1 is different from the Sulfolobus DinB proteins, which are members of cluster I. In addition to translesion DNA synthesis activity, MacDinB-1 synthesized unusually long products ( approximately 7.2 kb) in the presence of its cognate proliferating cell nuclear antigen (PCNA). The PCNA-interacting site in MacDinB-1 was identified by mutational analysis in a C-terminally located heptapeptide akin to a PIP (PCNA-interacting protein) box. In vitro assays from the present report suggested that MacDinB-1 works in an error-free mode to repair cyclobutane pyrimidine dimers. This study on a euryarchaeal DinB homolog provides important insights into the functional diversity of the family Y Pols, and the availability of a genetic system for this archaeon should allow subsequent elucidation of the physiological significance of this enzyme in M. acetivorans cells.
Lin
Li-Jung
LJ
Department of Animal Sciences, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
Yoshinaga
Aya
A
Lin
Yuyen
Y
Guzman
Claudia
C
Chen
Yi-Hsing
YH
Mei
Shou
S
Lagunas
Angelica M
AM
Koike
Satoshi
S
Iwai
Shigenori
S
Spies
M Ashley
MA
Nair
Satish K
SK
Mackie
Roderick I
RI
Ishino
Yoshizumi
Y
Cann
Isaac K O
IK
D000225:Adenine / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000302:isolation & purification; D004249:DNA Damage; D017931:DNA Primers / Q000378:metabolism; D019641:DNA, Archaeal / Q000096:biosynthesis; D004259:DNA-Directed DNA Polymerase / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D017020:Methanosarcina / Q000201:enzymology*; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics; D009154:Mutation / Q000235:genetics; D010802:Phylogeny; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D011485:Protein Binding; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D019843:Archaeal Proteins; D017931:DNA Primers; D019641:DNA, Archaeal; D018809:Proliferating Cell Nuclear Antigen; D004259:DNA-Directed DNA Polymerase; D000225:Adenine