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. 1991 Sep;173(17):5539–5545. doi: 10.1128/jb.173.17.5539-5545.1991

An essential iteron-binding protein required for plasmid R1162 replication induces localized melting within the origin at a specific site in AT-rich DNA.

Y J Kim 1, R J Meyer 1
PMCID: PMC208268  PMID: 1885530

Abstract

The R1162-encoded protein RepIB is essential for replication of the plasmid and binds specifically to iterons within the replicative origin. The protein causes the localized melting of DNA (determined by sensitivity to P1 nuclease) at a site within the AT-rich region of the origin, about 60 bp from the iteron binding sites and separated from them by a GC-rich tract. Point mutations have been isolated in the AT-rich DNA. These mutations interfere with origin activity and also prevent the protein-induced sensitivity to P1. A second-site suppressor of one of these mutations maps in the repIb gene and restores both origin function and sensitivity to P1. The results suggest a specific interaction between RepIB and origin DNA at a position distant from its primary binding site.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Amann E., Brosius J., Ptashne M. Vectors bearing a hybrid trp-lac promoter useful for regulated expression of cloned genes in Escherichia coli. Gene. 1983 Nov;25(2-3):167–178. doi: 10.1016/0378-1119(83)90222-6. [DOI] [PubMed] [Google Scholar]
  2. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  3. Bramhill D., Kornberg A. A model for initiation at origins of DNA replication. Cell. 1988 Sep 23;54(7):915–918. doi: 10.1016/0092-8674(88)90102-x. [DOI] [PubMed] [Google Scholar]
  4. Bramhill D., Kornberg A. Duplex opening by dnaA protein at novel sequences in initiation of replication at the origin of the E. coli chromosome. Cell. 1988 Mar 11;52(5):743–755. doi: 10.1016/0092-8674(88)90412-6. [DOI] [PubMed] [Google Scholar]
  5. Churchward G., Linder P., Caro L. The nucleotide sequence of replication and maintenance functions encoded by plasmid pSC101. Nucleic Acids Res. 1983 Aug 25;11(16):5645–5659. doi: 10.1093/nar/11.16.5645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Coggins L. W., McCluskey E. Characterization of the ColE1-Km plasmids pCR1 and pCR11 by electron microscope and restriction endonuclease mapping. Plasmid. 1979 Jul;2(3):446–453. doi: 10.1016/0147-619x(79)90028-3. [DOI] [PubMed] [Google Scholar]
  7. Froehlich B. J., Scott J. R. Second-site revertants of the P1 copN22 copy mutant. J Bacteriol. 1990 May;172(5):2762–2764. doi: 10.1128/jb.172.5.2762-2764.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fuller R. S., Funnell B. E., Kornberg A. The dnaA protein complex with the E. coli chromosomal replication origin (oriC) and other DNA sites. Cell. 1984 Oct;38(3):889–900. doi: 10.1016/0092-8674(84)90284-8. [DOI] [PubMed] [Google Scholar]
  9. Hershfield V., Boyer H. W., Yanofsky C., Lovett M. A., Helinski D. R. Plasmid ColEl as a molecular vehicle for cloning and amplification of DNA. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3455–3459. doi: 10.1073/pnas.71.9.3455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Honda Y., Sakai H., Komano T., Bagdasarian M. RepB' is required in trans for the two single-strand DNA initiation signals in oriV of plasmid RSF1010. Gene. 1989 Aug 1;80(1):155–159. doi: 10.1016/0378-1119(89)90261-8. [DOI] [PubMed] [Google Scholar]
  11. Kim K., Meyer R. J. Copy number of the broad host-range plasmid R1162 is determined by the amounts of essential plasmid-encoded proteins. J Mol Biol. 1985 Oct 20;185(4):755–767. doi: 10.1016/0022-2836(85)90060-9. [DOI] [PubMed] [Google Scholar]
  12. Kim Y. J., Lin L. S., Meyer R. J. Two domains at the origin are required for replication and maintenance of broad-host-range plasmid R1162. J Bacteriol. 1987 Dec;169(12):5870–5872. doi: 10.1128/jb.169.12.5870-5872.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kunkel T. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Proc Natl Acad Sci U S A. 1985 Jan;82(2):488–492. doi: 10.1073/pnas.82.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lin L. S., Meyer R. J. DNA synthesis is initiated at two positions within the origin of replication of plasmid R1162. Nucleic Acids Res. 1987 Oct 26;15(20):8319–8331. doi: 10.1093/nar/15.20.8319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lin L. S., Meyer R. J. Directly repeated, 20-bp sequence of plasmid R1162 DNA is required for replication, expression of incompatibility, and copy-number control. Plasmid. 1986 Jan;15(1):35–47. doi: 10.1016/0147-619x(86)90012-0. [DOI] [PubMed] [Google Scholar]
  16. Marko M. A., Chipperfield R., Birnboim H. C. A procedure for the large-scale isolation of highly purified plasmid DNA using alkaline extraction and binding to glass powder. Anal Biochem. 1982 Apr;121(2):382–387. doi: 10.1016/0003-2697(82)90497-3. [DOI] [PubMed] [Google Scholar]
  17. Meijer M., Beck E., Hansen F. G., Bergmans H. E., Messer W., von Meyenburg K., Schaller H. Nucleotide sequence of the origin of replication of the Escherichia coli K-12 chromosome. Proc Natl Acad Sci U S A. 1979 Feb;76(2):580–584. doi: 10.1073/pnas.76.2.580. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Miller J. H., Ganem D., Lu P., Schmitz A. Genetic studies of the lac repressor. I. Correlation of mutational sites with specific amino acid residues: construction of a colinear gene-protein map. J Mol Biol. 1977 Jan 15;109(2):275–298. doi: 10.1016/s0022-2836(77)80034-x. [DOI] [PubMed] [Google Scholar]
  19. Monk M., Kinross J. Conditional lethality of recA and recB derivatives of a strain of Escherichia coli K-12 with a temperature-sensitive deoxyribonucleic acid polymerase I. J Bacteriol. 1972 Mar;109(3):971–978. doi: 10.1128/jb.109.3.971-978.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Moore D. D., Denniston-Thompson K., Kruger K. E., Furth M. E., Williams B. G., Daniels D. L., Blattner F. R. Dissection and comparative anatomy of the origins of replication of lambdoid phages. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):155–163. doi: 10.1101/sqb.1979.043.01.022. [DOI] [PubMed] [Google Scholar]
  21. Scherzinger E., Bagdasarian M. M., Scholz P., Lurz R., Rückert B., Bagdasarian M. Replication of the broad host range plasmid RSF1010: requirement for three plasmid-encoded proteins. Proc Natl Acad Sci U S A. 1984 Feb;81(3):654–658. doi: 10.1073/pnas.81.3.654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schnos M., Zahn K., Inman R. B., Blattner F. R. Initiation protein induced helix destabilization at the lambda origin: a prepriming step in DNA replication. Cell. 1988 Feb 12;52(3):385–395. doi: 10.1016/s0092-8674(88)80031-x. [DOI] [PubMed] [Google Scholar]
  23. Stalker D. M., Thomas C. M., Helinski D. R. Nucleotide sequence of the region of the origin of replication of the broad host range plasmid RK2. Mol Gen Genet. 1981;181(1):8–12. doi: 10.1007/BF00338997. [DOI] [PubMed] [Google Scholar]
  24. Yung B. Y., Kornberg A. The dnaA initiator protein binds separate domains in the replication origin of Escherichia coli. J Biol Chem. 1989 Apr 15;264(11):6146–6150. [PubMed] [Google Scholar]
  25. Zhou H. S., Meyer R. J. Deletion of sites for initiation of DNA synthesis in the origin of broad host-range plasmid R1162. J Mol Biol. 1990 Aug 5;214(3):685–697. doi: 10.1016/0022-2836(90)90286-u. [DOI] [PubMed] [Google Scholar]

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