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J Biol Chem, Vol. 275, Issue 19, 14678-14683, May 12, 2000
From the Centro de Biología Molecular "Severo Ochoa"
(CSIC-UAM), Universidad Autónoma, Canto Blanco,
28049 Madrid, Spain
The linear genome of Bacillus
subtilis phage
Specific Recognition of Parental Terminal Protein by DNA
Polymerase for Initiation of Protein-primed DNA Replication*
,
29 has a protein covalently linked to the 5'
ends, called parental terminal protein (TP), and is replicated using a
free TP as primer. The initiation of phage 29 DNA replication
requires the formation of a DNA polymerase/TP complex that recognizes
the replication origins located at the genome ends. The DNA polymerase
catalyzes the formation of the initiation complex TP-dAMP, and
elongation proceeds coupled to strand displacement. The same mechanism
is used by the related phage Nf. However, DNA polymerase and TP from 29 do not initiate the replication of Nf TP-DNA. To address the question of the specificity of origin recognition, we took advantage of
the initiation reaction enhancement in the presence of
Mn2+, allowing us to detect initiation activity in
heterologous systems in which DNA polymerase, TP, and template TP-DNA
are not from the same phage. Initiation was selectively stimulated when
DNA polymerase and TP-DNA were from the same phage, strongly suggesting that specific recognition of origins is brought through an interaction between DNA polymerase and parental TP.
*
This work was supported by Research Grant 5R01 GM27242-20
from the National Institutes of Health, by Grant PB93-0173 from Dirección General de Investigación Científica y
Técnica, by Grant ERBFMX CT97 0125 from European Union, and by an
Institutional grant from Fundación Ramón Areces.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Predoctoral fellow of the Gobierno Vasco.
§
To whom correspondence should be addressed. Tel.: 34-91-3978435;
Fax: 34-91-3978490; E-mail: msalas@cbm.uam.es.
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