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. 1988 Jun;7(6):1881–1888. doi: 10.1002/j.1460-2075.1988.tb03021.x

Recognition of the P1 plasmid centromere analog involves binding of the ParB protein and is modified by a specific host factor.

M A Davis 1, S J Austin 1
PMCID: PMC457181  PMID: 3049080

Abstract

The P1 plasmid partition system is responsible for segregation of daughter plasmids during division of the Escherichia coli host cell. The P1-encoded elements consist of two essential proteins, ParA and ParB, and the cis-acting incB region. The incB region determines partition-mediated incompatibility and contains the centromere-like site parS. We have isolated and purified the two proteins. ParB binds specifically to the incB region in vitro. DNase I footprinting assays place a strong binding site over the 35-bp parS sequence previously shown to be sufficient for partition when the Par proteins are supplied in trans. A weaker site lies within the incB region in sequences that are important for specifying incompatibility, but are not essential for partition. Gel band retardation assays show that a host factor binds specifically to the incB sequence. The factor strongly stimulates binding of ParB. Cutting the region at a site between the two ParB binding sites yields two fragments that can bind ParB but not host factor. Thus, information for host-factor binding lies in the region determining the specificity of plasmid incompatibility. The roles of parB and the host factor in partition and the specificity of plasmid incompatibility are discussed.

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Selected References

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  1. Abeles A. L., Friedman S. A., Austin S. J. Partition of unit-copy miniplasmids to daughter cells. III. The DNA sequence and functional organization of the P1 partition region. J Mol Biol. 1985 Sep 20;185(2):261–272. doi: 10.1016/0022-2836(85)90402-4. [DOI] [PubMed] [Google Scholar]
  2. Abeles A. L. P1 plasmid replication. Purification and DNA-binding activity of the replication protein RepA. J Biol Chem. 1986 Mar 15;261(8):3548–3555. [PubMed] [Google Scholar]
  3. Abremski K., Hoess R. Escherichia coli plasmid vectors for high-level regulated expression of the bacteriophage lambda xis gene product. Gene. 1983 Nov;25(1):49–58. doi: 10.1016/0378-1119(83)90166-x. [DOI] [PubMed] [Google Scholar]
  4. Austin S., Abeles A. Partition of unit-copy miniplasmids to daughter cells. I. P1 and F miniplasmids contain discrete, interchangeable sequences sufficient to promote equipartition. J Mol Biol. 1983 Sep 15;169(2):353–372. doi: 10.1016/s0022-2836(83)80055-2. [DOI] [PubMed] [Google Scholar]
  5. Austin S., Abeles A. Partition of unit-copy miniplasmids to daughter cells. II. The partition region of miniplasmid P1 encodes an essential protein and a centromere-like site at which it acts. J Mol Biol. 1983 Sep 15;169(2):373–387. doi: 10.1016/s0022-2836(83)80056-4. [DOI] [PubMed] [Google Scholar]
  6. Austin S., Friedman S., Ludtke D. Partition functions of unit-copy plasmids can stabilize the maintenance of plasmid pBR322 at low copy number. J Bacteriol. 1986 Nov;168(2):1010–1013. doi: 10.1128/jb.168.2.1010-1013.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Austin S., Hart F., Abeles A., Sternberg N. Genetic and physical map of a P1 miniplasmid. J Bacteriol. 1982 Oct;152(1):63–71. doi: 10.1128/jb.152.1.63-71.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fried M., Crothers D. M. Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res. 1981 Dec 11;9(23):6505–6525. doi: 10.1093/nar/9.23.6505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Galas D. J., Schmitz A. DNAse footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res. 1978 Sep;5(9):3157–3170. doi: 10.1093/nar/5.9.3157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Garner M. M., Revzin A. A gel electrophoresis method for quantifying the binding of proteins to specific DNA regions: application to components of the Escherichia coli lactose operon regulatory system. Nucleic Acids Res. 1981 Jul 10;9(13):3047–3060. doi: 10.1093/nar/9.13.3047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Martin K. A., Friedman S. A., Austin S. J. Partition site of the P1 plasmid. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8544–8547. doi: 10.1073/pnas.84.23.8544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  13. Novick R. P., Hoppensteadt F. C. On plasmid incompatibility. Plasmid. 1978 Sep;1(4):421–434. doi: 10.1016/0147-619x(78)90001-x. [DOI] [PubMed] [Google Scholar]
  14. Sternberg N., Weisberg R. Packaging of prophage and host DNA by coliphage lambda. Nature. 1975 Jul 10;256(5513):97–103. doi: 10.1038/256097a0. [DOI] [PubMed] [Google Scholar]
  15. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Zuber M., Patterson T. A., Court D. L. Analysis of nutR, a site required for transcription antitermination in phage lambda. Proc Natl Acad Sci U S A. 1987 Jul;84(13):4514–4518. doi: 10.1073/pnas.84.13.4514. [DOI] [PMC free article] [PubMed] [Google Scholar]

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