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DnaB proteolysis in vivo regulates oligomerization and its localization at oriC in Bacillus subtilis

Grainger, William H.; Machón, Cristina; Scott, David J.; Soultanas, Panos

Authors

William H. Grainger

Cristina Machón

David J. Scott



Abstract

Initiation of bacterial DNA replication at oriC is mediated by primosomal proteins that act cooperatively to melt an AT-rich region where the replicative helicase is loaded prior to the assembly of the replication fork. In Bacillus subtilis, the dnaD, dnaB and dnaI genes are essential for initiation of DNA replication. We established that their mRNAs are maintained in fast growing asynchronous cultures. DnaB is truncated at its C-terminus in a growth phase-dependent manner. Proteolysis is confined to cytosolic, not to membrane-associated DnaB, and affects oligomerization. Truncated DnaB is depleted at the oriC relative to the native protein. We propose that DNA-induced oligomerization is essential for its action at oriC and proteolysis regulates its localization at oriC. We show that DnaB has two separate ssDNA-binding sites one located within residues 1-300 and another between residues 365-428, and a dsDNA-binding site within residues 365-428. Tetramerization of DnaB is mediated within residues 1-300, and DNA-dependent oligomerization within residues 365-428. Finally, we show that association of DnaB with the oriC is asymmetric and extensive. It encompasses an area from the middle of dnaA to the end of yaaA that includes the AT-rich region melted during the initiation stage of DNA replication.

Citation

Grainger, W. H., Machón, C., Scott, D. J., & Soultanas, P. (2010). DnaB proteolysis in vivo regulates oligomerization and its localization at oriC in Bacillus subtilis. Nucleic Acids Research, 38(9), doi:10.1093/nar/gkp1236

Journal Article Type Article
Publication Date May 1, 2010
Deposit Date Jun 2, 2010
Publicly Available Date Jun 2, 2010
Journal Nucleic Acids Research
Print ISSN 0305-1048
Electronic ISSN 0305-1048
Publisher Oxford University Press (OUP)
Peer Reviewed Peer Reviewed
Volume 38
Issue 9
DOI https://doi.org/10.1093/nar/gkp1236
Public URL http://eprints.nottingham.ac.uk/id/eprint/1311
Publisher URL http://nar.oxfordjournals.org/cgi/content/full/38/9/2851
Copyright Statement Copyright information regarding this work can be found at the following address: http://eprints.nottingham.ac.uk/end_user_agreement.pdf

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://eprints.nottingham.ac.uk/end_user_agreement.pdf





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