Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1983 Jun;154(3):1184–1194. doi: 10.1128/jb.154.3.1184-1194.1983

Site-specific in vitro binding of plasmid pUB110 to Bacillus subtilis membrane fraction.

T Tanaka, N Sueoka
PMCID: PMC217590  PMID: 6406425

Abstract

The in vitro membrane binding of pSL103, a composite plasmid consisting of Staphylococcus aureus plasmid pUB110 and a Bacillus pumilus trpC+ DNA fragment, to the Bacillus subtilis membrane fraction was studied with a total lysate of B. subtilis cells. The binding reaction required a heat treatment at 45 degrees C and had an optimum KCl concentration of 60 mM. Nonradioactive pSL103, but not Escherichia coli plasmid pACYC184, competed with 3H-labeled pSL103 for binding to the membrane. By the use of 32P-labeled restriction fragments of pSL103 and pUB110, it has been found that only the pUB110 portion of pSL103 binds to the membrane and that there are four specific regions in pUB110 which bind to the membrane. Two of the four binding regions flank the replication origin. This in vitro binding was high-salt sensitive and apparently independent of the configurations of the plasmid. We have previously shown that the functional product of the initiation gene dna-1 is required in vivo both for replication initiation and the binding of a DNA region near the replication origin to the membrane. Unlike in vivo binding, which is high-salt resistant and dependent on the product of dna-1 gene (type-I binding), the in vitro binding reported in this paper was high-salt sensitive and independent of the dna-1 gene product (type-II binding).

Full text

PDF
1194

Images in this article

1188Image
on p.1188
1189Image
on p.1189
1190Image
on p.1190
1192Image
on p.1192

Selected References

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

  1. Alwine J. C., Kemp D. J., Parker B. A., Reiser J., Renart J., Stark G. R., Wahl G. M. Detection of specific RNAs or specific fragments of DNA by fractionation in gels and transfer to diazobenzyloxymethyl paper. Methods Enzymol. 1979;68:220–242. doi: 10.1016/0076-6879(79)68017-5. [DOI] [PubMed] [Google Scholar]
  2. Craine B. L., Rupert C. S. Deoxyribonucleic acid-membrane interactions near the origin of replication and initiation of deoxyribonucleic acid synthesis in Escherichia coli. J Bacteriol. 1979 Feb;137(2):740–745. doi: 10.1128/jb.137.2.740-745.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Diaz R., Nordström K., Staudenbauer W. L. Plasmid R1 DNA replication dependent on protein synthesis in cell-free extracts of E. coli. Nature. 1981 Jan 22;289(5795):326–328. doi: 10.1038/289326a0. [DOI] [PubMed] [Google Scholar]
  4. Fielding P., Fox C. F. Evidence for stable attachment of DNA to membrane at the replication origin of Escherichia coli. Biochem Biophys Res Commun. 1970 Oct 9;41(1):157–162. doi: 10.1016/0006-291x(70)90482-1. [DOI] [PubMed] [Google Scholar]
  5. Firshein W. The DNA-membrane fraction of Pneumococcus contains a DNA replication complex. J Mol Biol. 1972 Oct 14;70(3):383–397. doi: 10.1016/0022-2836(72)90547-5. [DOI] [PubMed] [Google Scholar]
  6. Fuller R. S., Kaguni J. M., Kornberg A. Enzymatic replication of the origin of the Escherichia coli chromosome. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7370–7374. doi: 10.1073/pnas.78.12.7370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gryczan T. J., Contente S., Dubnau D. Characterization of Staphylococcus aureus plasmids introduced by transformation into Bacillus subtilis. J Bacteriol. 1978 Apr;134(1):318–329. doi: 10.1128/jb.134.1.318-329.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Imada S., Carroll L. E., Sueoka N. Genetic mapping of a group of temperature-sensitive dna initiation mutants in Bacillus subtilis. Genetics. 1980 Apr;94(4):809–823. doi: 10.1093/genetics/94.4.809. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Inuzuka M., Helinski D. R. Requirement of a plasmid-encoded protein for replication in vitro of plasmid R6K. Proc Natl Acad Sci U S A. 1978 Nov;75(11):5381–5385. doi: 10.1073/pnas.75.11.5381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Karamata D., Gross J. D. Isolation and genetic analysis of temperature-sensitive mutants of B. subtilis defective in DNA synthesis. Mol Gen Genet. 1970;108(3):277–287. doi: 10.1007/BF00283358. [DOI] [PubMed] [Google Scholar]
  11. Keggins K. M., Lovett P. S., Duvall E. J. Molecular cloning of genetically active fragments of Bacillus DNA in Bacillus subtilis and properties of the vector plasmid pUB110. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1423–1427. doi: 10.1073/pnas.75.3.1423. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Keggins K. M., Lovett P. S., Marrero R., Hoch S. O. Insertional inactivation of trpC in cloned Bacillus trp segments: evidence for a polar effect on trpF. J Bacteriol. 1979 Sep;139(3):1001–1006. doi: 10.1128/jb.139.3.1001-1006.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Korn R., Winston S., Tanaka T., Sueoka N. Specific in vitro binding of a plasmid to a membrane fraction of Bacillus subtilis. Proc Natl Acad Sci U S A. 1983 Jan;80(2):574–578. doi: 10.1073/pnas.80.2.574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  15. Meacock P. A., Cohen S. N. Partitioning of bacterial plasmids during cell division: a cis-acting locus that accomplishes stable plasmid inheritance. Cell. 1980 Jun;20(2):529–542. doi: 10.1016/0092-8674(80)90639-x. [DOI] [PubMed] [Google Scholar]
  16. Nagai K., Hendrickson W., Balakrishnan R., Yamaki H., Boyd D., Schaechter M. Isolation of a replication origin complex from Escherichia coli. Proc Natl Acad Sci U S A. 1980 Jan;77(1):262–266. doi: 10.1073/pnas.77.1.262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Parker D. L., Glaser D. A. Chromosomal sites of DNA-membrane attachment in Escherichia coli. J Mol Biol. 1974 Aug 5;87(2):153–168. doi: 10.1016/0022-2836(74)90140-5. [DOI] [PubMed] [Google Scholar]
  18. RYTER A., JACOB F. ETUDE AU MICROSCOPE 'ELECTRONIQUE DES RELATIONS ENTRE M'ESOSOMES ET NOYAUX CHEZ BACILLUS SUBTILIS. C R Hebd Seances Acad Sci. 1963 Nov 13;257:3060–3063. [PubMed] [Google Scholar]
  19. Rownd R. Replication of a bacterial episome under relaxed control. J Mol Biol. 1969 Sep 28;44(3):387–402. doi: 10.1016/0022-2836(69)90368-4. [DOI] [PubMed] [Google Scholar]
  20. Scheer-Abramowitz J., Gryczan T. J., Dubnau D. Origin and mode of replication of plasmids pE194 and pUB110. Plasmid. 1981 Jul;6(1):67–77. doi: 10.1016/0147-619x(81)90054-8. [DOI] [PubMed] [Google Scholar]
  21. Smith D. W., Hanawalt P. C. Properties of the growing point region in the bacterial chromosome. Biochim Biophys Acta. 1967 Dec 19;149(2):519–531. doi: 10.1016/0005-2787(67)90180-3. [DOI] [PubMed] [Google Scholar]
  22. Snyder R. W., Young F. E. Association between the chromosome and the cytoplasmic membrane in Bacillus subtilis. Biochem Biophys Res Commun. 1969 May 8;35(3):354–362. doi: 10.1016/0006-291x(69)90506-3. [DOI] [PubMed] [Google Scholar]
  23. Sueoka N., Hammers J. M. Isolation of DNA-membrane complex in Bacillus subtilis. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4787–4791. doi: 10.1073/pnas.71.12.4787. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sueoka N., Quinn W. G. Membrane attachment of the chromosome replication origin in Bacillus subtilis. Cold Spring Harb Symp Quant Biol. 1968;33:695–705. doi: 10.1101/sqb.1968.033.01.078. [DOI] [PubMed] [Google Scholar]
  25. Tanaka T., Kawano N. Cloning vehicles for the homologous Bacillus subtilis host-vector system. Gene. 1980 Jul;10(2):131–136. doi: 10.1016/0378-1119(80)90130-4. [DOI] [PubMed] [Google Scholar]
  26. Tomizawa J., Itoh T., Selzer G., Som T. Inhibition of ColE1 RNA primer formation by a plasmid-specified small RNA. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1421–1425. doi: 10.1073/pnas.78.3.1421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. White K., Sueoka N. Temperature-sensitive DNA synthesis mutants of Bacillus subtilis--appendix: theory of density transfer for symmetric chromosome replication. Genetics. 1973 Feb;73(2):185–214. doi: 10.1093/genetics/73.2.185. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Winston S., Sueoka N. DNA-membrane association is necessary for initiation of chromosomal and plasmid replication in Bacillus subtilis. Proc Natl Acad Sci U S A. 1980 May;77(5):2834–2838. doi: 10.1073/pnas.77.5.2834. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Yamaguchi K., Yoshikawa H. Association of the replication terminus of the Bacillus subtilis chromosome to the cell membrane. J Bacteriol. 1975 Nov;124(2):1030–1033. doi: 10.1128/jb.124.2.1030-1033.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Yamaguchi K., Yoshikawa H. Topography of chromosome membrane junction in Bacillus subtilis. Nat New Biol. 1973 Aug 15;244(137):204–206. doi: 10.1038/newbio244204a0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES