Affiliation:
1. Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709
2. D. I. Ivanovsky Institute of Virology, Russian Academy of Medical Science, Moscow, 123098, Russia
Abstract
ABSTRACT
The θ subunit (
holE
gene product) of
Escherichia coli
DNA polymerase (Pol) III holoenzyme is a tightly bound component of the polymerase core. Within the core (α-ε-θ), the α and ε subunits carry the DNA polymerase and 3′ proofreading functions, respectively, while the precise function of θ is unclear.
holE
homologs are present in genomes of other enterobacteriae, suggestive of a conserved function. Putative homologs have also been found in the genomes of bacteriophage P1 and of certain conjugative plasmids. The presence of these homologs is of interest, because these genomes are fully dependent on the host replication machinery and contribute few, if any, replication factors themselves. To study the role of these θ homologs, we have constructed an
E. coli
strain in which
holE
is replaced by the P1 homolog,
hot
. We show that
hot
is capable of substituting for
holE
when it is assayed for its antimutagenic action on the proofreading-impaired
dnaQ49
mutator, which carries a temperature-sensitive ε subunit. The ability of
hot
to substitute for
holE
was also observed with other, although not all,
dnaQ
mutator alleles tested. The data suggest that the P1
hot
gene product can substitute for the θ subunit and is likely incorporated in the Pol III complex. We also show that overexpression of either θ or Hot further suppresses the
dnaQ49
mutator phenotype. This suggests that the complexing of
dnaQ49
-ε with θ is rate limiting for its ability to proofread DNA replication errors. The possible role of
hot
for bacteriophage P1 is discussed.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Reference52 articles.
1. Akman, L., A. Yamashita, H. Watanabe, K. Oshima, T. Shiba, M. Hattori, and S. Aksoy. 2002. Genome sequence of the endocellular obligate symbiont of tsetse flies, Wigglesworthia glossinidia. Nat. Genet.32:402-407.
2. Carter, J. R., M. A. Franden, R. Aebersold, D. R. Kim, and C. S. McHenry. 1993. Isolation, sequencing and overexpression of the gene encoding the θ subunit of DNA polymerase III holoenzyme. Nucleic Acids Res.21:3281-3286.
3. DeRose, E. F., T. Darden, S. Harvey, S. Gabel, F. W. Perrino, R. M. Schaaper, and R. E. London. 2003. Elucidation of the ε-θ interface of Escherichia coli DNA polymerase III by NMR spectroscopy. Biochemistry42:3635-3644.
4. DeRose, E. F., T. W. Kirby, G. A. Mueller, A. K. Chikova, R. M. Schaaper, and R. E. London. 2004. Phage like it HOT: solution structure of the bacteriophage P1-encoded HOT protein, a homolog of the θ subunit of E. coli DNA polymerase III. Structure12:2221-2231.
5. DeRose, E. F., D. Li, T. Darden, S. Harvey, F. W. Perrino, R. M. Schaaper, and R. E. London. 2002. Model for the catalytic domain of the proofreading epsilon subunit of Escherichia coli DNA polymerase III based on NMR structural data. Biochemistry41:94-110.
Cited by
19 articles.
订阅此论文施引文献
订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献