Functional interplay of DnaE polymerase, DnaG primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis
Bacillus subtilis has two replicative DNA polymerases. PolC is a processive high-fidelity replicative polymerase, while the error-prone DnaEBs extends RNA primers before hand-off to PolC at the lagging strand. We show that DnaEBs interacts with the replicative helicase DnaC and primase DnaG in...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Published: |
Oxford Journals
2013
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| Online Access: | https://eprints.nottingham.ac.uk/2043/ |
| _version_ | 1848790711438147584 |
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| author | Rannou, Olivier Le Chatelier, Emanuelle Larson, Marilynn A. Nouri, Hamid Dalmais, Bérengère Laughton, Charles Laurent, Jannière Soultanas, Panos |
| author_facet | Rannou, Olivier Le Chatelier, Emanuelle Larson, Marilynn A. Nouri, Hamid Dalmais, Bérengère Laughton, Charles Laurent, Jannière Soultanas, Panos |
| author_sort | Rannou, Olivier |
| building | Nottingham Research Data Repository |
| collection | Online Access |
| description | Bacillus subtilis has two replicative DNA polymerases.
PolC is a processive high-fidelity replicative
polymerase, while the error-prone DnaEBs extends
RNA primers before hand-off to PolC at the lagging
strand. We show that DnaEBs interacts with the replicative
helicase DnaC and primase DnaG in a
ternary complex. We characterize their activities
and analyse the functional significance of their interactions
using primase, helicase and primer extension
assays, and a ‘stripped down’ reconstituted
coupled assay to investigate the coordinated displacement
of the parental duplex DNA at a replication
fork, synthesis of RNA primers along the
lagging strand and hand-off to DnaEBs. The DnaG–
DnaEBs hand-off takes place after de novo polymerization
of only two ribonucleotides by DnaG, and
does not require other replication proteins.
Furthermore, the fidelity of DnaEBs is improved by
DnaC and DnaG, likely via allosteric effects induced
by direct protein–protein interactions that lower the
efficiency of nucleotide mis-incorporations and/or
the efficiency of extension of mis-aligned primers
in the catalytic site of DnaEBs. We conclude that
de novo RNA primer synthesis by DnaG and initial
primer extension by DnaEBs are carried out by a
lagging strand–specific subcomplex comprising
DnaG, DnaEBs and DnaC, which stimulates chromosomal
replication with enhanced fidelity. |
| first_indexed | 2025-11-14T18:16:57Z |
| format | Article |
| id | nottingham-2043 |
| institution | University of Nottingham Malaysia Campus |
| institution_category | Local University |
| last_indexed | 2025-11-14T18:16:57Z |
| publishDate | 2013 |
| publisher | Oxford Journals |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | nottingham-20432020-05-04T16:36:10Z https://eprints.nottingham.ac.uk/2043/ Functional interplay of DnaE polymerase, DnaG primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis Rannou, Olivier Le Chatelier, Emanuelle Larson, Marilynn A. Nouri, Hamid Dalmais, Bérengère Laughton, Charles Laurent, Jannière Soultanas, Panos Bacillus subtilis has two replicative DNA polymerases. PolC is a processive high-fidelity replicative polymerase, while the error-prone DnaEBs extends RNA primers before hand-off to PolC at the lagging strand. We show that DnaEBs interacts with the replicative helicase DnaC and primase DnaG in a ternary complex. We characterize their activities and analyse the functional significance of their interactions using primase, helicase and primer extension assays, and a ‘stripped down’ reconstituted coupled assay to investigate the coordinated displacement of the parental duplex DNA at a replication fork, synthesis of RNA primers along the lagging strand and hand-off to DnaEBs. The DnaG– DnaEBs hand-off takes place after de novo polymerization of only two ribonucleotides by DnaG, and does not require other replication proteins. Furthermore, the fidelity of DnaEBs is improved by DnaC and DnaG, likely via allosteric effects induced by direct protein–protein interactions that lower the efficiency of nucleotide mis-incorporations and/or the efficiency of extension of mis-aligned primers in the catalytic site of DnaEBs. We conclude that de novo RNA primer synthesis by DnaG and initial primer extension by DnaEBs are carried out by a lagging strand–specific subcomplex comprising DnaG, DnaEBs and DnaC, which stimulates chromosomal replication with enhanced fidelity. Oxford Journals 2013-05-01 Article PeerReviewed Rannou, Olivier, Le Chatelier, Emanuelle, Larson, Marilynn A., Nouri, Hamid, Dalmais, Bérengère, Laughton, Charles, Laurent, Jannière and Soultanas, Panos (2013) Functional interplay of DnaE polymerase, DnaG primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis. Nucleic Acids Research, 41 (10). pp. 5303-5320. ISSN 0305-1048 http://www.ncbi.nlm.nih.gov/pubmed/23563155 doi:10.1093/nar/gkt207 doi:10.1093/nar/gkt207 |
| spellingShingle | Rannou, Olivier Le Chatelier, Emanuelle Larson, Marilynn A. Nouri, Hamid Dalmais, Bérengère Laughton, Charles Laurent, Jannière Soultanas, Panos Functional interplay of DnaE polymerase, DnaG primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis |
| title | Functional interplay of DnaE polymerase, DnaG
primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis |
| title_full | Functional interplay of DnaE polymerase, DnaG
primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis |
| title_fullStr | Functional interplay of DnaE polymerase, DnaG
primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis |
| title_full_unstemmed | Functional interplay of DnaE polymerase, DnaG
primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis |
| title_short | Functional interplay of DnaE polymerase, DnaG
primase and DnaC helicase within a ternary complex, and primase to polymerase hand-off during lagging strand DNA replication in Bacillus subtilis |
| title_sort | functional interplay of dnae polymerase, dnag
primase and dnac helicase within a ternary complex, and primase to polymerase hand-off during lagging strand dna replication in bacillus subtilis |
| url | https://eprints.nottingham.ac.uk/2043/ https://eprints.nottingham.ac.uk/2043/ https://eprints.nottingham.ac.uk/2043/ |