Positioning and the specific sequence of each 13-mer motif are critical for activity of the plasmid RK2 replication origin.
Identifieur interne : 002E90 ( Main/Exploration ); précédent : 002E89; suivant : 002E91Positioning and the specific sequence of each 13-mer motif are critical for activity of the plasmid RK2 replication origin.
Auteurs : Lukasz Kowalczyk [Pologne] ; Magdalena Rajewska ; Igor KoniecznySource :
- Molecular microbiology [ 0950-382X ] ; 2005.
Descripteurs français
- KwdFr :
- ADN bactérien (génétique), ADN bactérien (métabolisme), Données de séquences moléculaires, Escherichia coli (génétique), Mutation, Origine de réplication (génétique), Origine de réplication (physiologie), Plasmides (génétique), Pseudomonas aeruginosa (génétique), Réplication de l'ADN, Séquence nucléotidique.
- MESH :
- génétique : ADN bactérien, Escherichia coli, Origine de réplication, Plasmides, Pseudomonas aeruginosa.
- métabolisme : ADN bactérien.
- physiologie : Origine de réplication.
- Données de séquences moléculaires, Mutation, Réplication de l'ADN, Séquence nucléotidique.
English descriptors
- KwdEn :
- MESH :
- chemical , genetics : DNA, Bacterial.
- chemical , metabolism : DNA, Bacterial.
- genetics : Escherichia coli, Plasmids, Pseudomonas aeruginosa, Replication Origin.
- physiology : Replication Origin.
- Base Sequence, DNA Replication, Molecular Sequence Data, Mutation.
Abstract
The minimal replication origin of the broad-host-range plasmid RK2, oriV, contains five iterons which are binding sites for the plasmid-encoded replication initiation protein TrfA, four DnaA boxes, which bind the host DnaA protein, and an AT-rich region containing four 13-mer sequences. In this study, 26 mutants with altered sequence and/or spacing of 13-mer motifs have been constructed and analysed for replication activity in vivo and in vitro. The data show that the replacement of oriV 13-mers by similar but not identical 13-mer sequences from Escherichia coli oriC inactivates the origin. In addition, interchanging the positions of the oriV 13-mers results in greatly reduced activity. Mutants with T/A substitutions are also inactive. Furthermore, introduction of single-nucleotide substitutions demonstrates very restricted sequence requirements depending on the 13-mer position. Only two of the mutants are host specific, functional in Pseudomonas aeruginosa but not in E. coli. Our experiments demonstrate considerable complexity in the plasmid AT-rich region architecture required for functionality. It is evident that low internal stability of this region is not the only feature contributing to origin activity. Our studies suggest a requirement for sequence-specific protein interactions within the 13-mers during assembly of replication complexes at the plasmid origin.
DOI: 10.1111/j.1365-2958.2005.04770.x
PubMed: 16102011
Affiliations:
Links toward previous steps (curation, corpus...)
- to stream PubMed, to step Corpus: 002317
- to stream PubMed, to step Curation: 002317
- to stream PubMed, to step Checkpoint: 002188
- to stream Ncbi, to step Merge: 000362
- to stream Ncbi, to step Curation: 000362
- to stream Ncbi, to step Checkpoint: 000362
- to stream Main, to step Merge: 002F20
- to stream Main, to step Curation: 002E90
Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Positioning and the specific sequence of each 13-mer motif are critical for activity of the plasmid RK2 replication origin.</title><author><name sortKey="Kowalczyk, Lukasz" sort="Kowalczyk, Lukasz" uniqKey="Kowalczyk L" first="Lukasz" last="Kowalczyk">Lukasz Kowalczyk</name><affiliation wicri:level="1"><nlm:affiliation>Department of Molecular and Cellular Biology, Intercollegiate Faculty of Biotechnology, University of Gdansk, ul. Kladki 24, 80-822 Gdansk, Poland.</nlm:affiliation><country xml:lang="fr">Pologne</country><wicri:regionArea>Department of Molecular and Cellular Biology, Intercollegiate Faculty of Biotechnology, University of Gdansk, ul. Kladki 24, 80-822 Gdansk</wicri:regionArea><wicri:noRegion>80-822 Gdansk</wicri:noRegion></affiliation></author><author><name sortKey="Rajewska, Magdalena" sort="Rajewska, Magdalena" uniqKey="Rajewska M" first="Magdalena" last="Rajewska">Magdalena Rajewska</name></author><author><name sortKey="Konieczny, Igor" sort="Konieczny, Igor" uniqKey="Konieczny I" first="Igor" last="Konieczny">Igor Konieczny</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2005">2005</date><idno type="RBID">pubmed:16102011</idno><idno type="pmid">16102011</idno><idno type="doi">10.1111/j.1365-2958.2005.04770.x</idno><idno type="wicri:Area/PubMed/Corpus">002317</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">002317</idno><idno type="wicri:Area/PubMed/Curation">002317</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">002317</idno><idno type="wicri:Area/PubMed/Checkpoint">002188</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">002188</idno><idno type="wicri:Area/Ncbi/Merge">000362</idno><idno type="wicri:Area/Ncbi/Curation">000362</idno><idno type="wicri:Area/Ncbi/Checkpoint">000362</idno><idno type="wicri:doubleKey">0950-382X:2005:Kowalczyk L:positioning:and:the</idno><idno type="wicri:Area/Main/Merge">002F20</idno><idno type="wicri:Area/Main/Curation">002E90</idno><idno type="wicri:Area/Main/Exploration">002E90</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Positioning and the specific sequence of each 13-mer motif are critical for activity of the plasmid RK2 replication origin.</title><author><name sortKey="Kowalczyk, Lukasz" sort="Kowalczyk, Lukasz" uniqKey="Kowalczyk L" first="Lukasz" last="Kowalczyk">Lukasz Kowalczyk</name><affiliation wicri:level="1"><nlm:affiliation>Department of Molecular and Cellular Biology, Intercollegiate Faculty of Biotechnology, University of Gdansk, ul. Kladki 24, 80-822 Gdansk, Poland.</nlm:affiliation><country xml:lang="fr">Pologne</country><wicri:regionArea>Department of Molecular and Cellular Biology, Intercollegiate Faculty of Biotechnology, University of Gdansk, ul. Kladki 24, 80-822 Gdansk</wicri:regionArea><wicri:noRegion>80-822 Gdansk</wicri:noRegion></affiliation></author><author><name sortKey="Rajewska, Magdalena" sort="Rajewska, Magdalena" uniqKey="Rajewska M" first="Magdalena" last="Rajewska">Magdalena Rajewska</name></author><author><name sortKey="Konieczny, Igor" sort="Konieczny, Igor" uniqKey="Konieczny I" first="Igor" last="Konieczny">Igor Konieczny</name></author></analytic><series><title level="j">Molecular microbiology</title><idno type="ISSN">0950-382X</idno><imprint><date when="2005" type="published">2005</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Base Sequence</term><term>DNA Replication</term><term>DNA, Bacterial (genetics)</term><term>DNA, Bacterial (metabolism)</term><term>Escherichia coli (genetics)</term><term>Molecular Sequence Data</term><term>Mutation</term><term>Plasmids (genetics)</term><term>Pseudomonas aeruginosa (genetics)</term><term>Replication Origin (genetics)</term><term>Replication Origin (physiology)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ADN bactérien (génétique)</term><term>ADN bactérien (métabolisme)</term><term>Données de séquences moléculaires</term><term>Escherichia coli (génétique)</term><term>Mutation</term><term>Origine de réplication (génétique)</term><term>Origine de réplication (physiologie)</term><term>Plasmides (génétique)</term><term>Pseudomonas aeruginosa (génétique)</term><term>Réplication de l'ADN</term><term>Séquence nucléotidique</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>DNA, Bacterial</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>DNA, Bacterial</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Escherichia coli</term><term>Plasmids</term><term>Pseudomonas aeruginosa</term><term>Replication Origin</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ADN bactérien</term><term>Escherichia coli</term><term>Origine de réplication</term><term>Plasmides</term><term>Pseudomonas aeruginosa</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>ADN bactérien</term></keywords><keywords scheme="MESH" qualifier="physiologie" xml:lang="fr"><term>Origine de réplication</term></keywords><keywords scheme="MESH" qualifier="physiology" xml:lang="en"><term>Replication Origin</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Base Sequence</term><term>DNA Replication</term><term>Molecular Sequence Data</term><term>Mutation</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Données de séquences moléculaires</term><term>Mutation</term><term>Réplication de l'ADN</term><term>Séquence nucléotidique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The minimal replication origin of the broad-host-range plasmid RK2, oriV, contains five iterons which are binding sites for the plasmid-encoded replication initiation protein TrfA, four DnaA boxes, which bind the host DnaA protein, and an AT-rich region containing four 13-mer sequences. In this study, 26 mutants with altered sequence and/or spacing of 13-mer motifs have been constructed and analysed for replication activity in vivo and in vitro. The data show that the replacement of oriV 13-mers by similar but not identical 13-mer sequences from Escherichia coli oriC inactivates the origin. In addition, interchanging the positions of the oriV 13-mers results in greatly reduced activity. Mutants with T/A substitutions are also inactive. Furthermore, introduction of single-nucleotide substitutions demonstrates very restricted sequence requirements depending on the 13-mer position. Only two of the mutants are host specific, functional in Pseudomonas aeruginosa but not in E. coli. Our experiments demonstrate considerable complexity in the plasmid AT-rich region architecture required for functionality. It is evident that low internal stability of this region is not the only feature contributing to origin activity. Our studies suggest a requirement for sequence-specific protein interactions within the 13-mers during assembly of replication complexes at the plasmid origin.</div></front></TEI><affiliations><list><country><li>Pologne</li></country></list><tree><noCountry><name sortKey="Konieczny, Igor" sort="Konieczny, Igor" uniqKey="Konieczny I" first="Igor" last="Konieczny">Igor Konieczny</name><name sortKey="Rajewska, Magdalena" sort="Rajewska, Magdalena" uniqKey="Rajewska M" first="Magdalena" last="Rajewska">Magdalena Rajewska</name></noCountry><country name="Pologne"><noRegion><name sortKey="Kowalczyk, Lukasz" sort="Kowalczyk, Lukasz" uniqKey="Kowalczyk L" first="Lukasz" last="Kowalczyk">Lukasz Kowalczyk</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Sante/explor/MersV1/Data/Main/Exploration
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 002E90 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd -nk 002E90 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Sante
|area= MersV1
|flux= Main
|étape= Exploration
|type= RBID
|clé= pubmed:16102011
|texte= Positioning and the specific sequence of each 13-mer motif are critical for activity of the plasmid RK2 replication origin.
}}
Pour générer des pages wiki
HfdIndexSelect -h $EXPLOR_AREA/Data/Main/Exploration/RBID.i -Sk "pubmed:16102011" \
| HfdSelect -Kh $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd \
| NlmPubMed2Wicri -a MersV1
| This area was generated with Dilib version V0.6.33. |  |