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. 1989 May;171(5):2466–2473. doi: 10.1128/jb.171.5.2466-2473.1989

The ssb gene of plasmid ColIb-P9.

C J Howland 1, C E Rees 1, P T Barth 1, B M Wilkins 1
PMCID: PMC209922  PMID: 2651402

Abstract

The IncI1 plasmid ColIb-P9 was found to carry a single-stranded DNA-binding (SSB) protein gene (ssb) that maps about 11 kilobase pairs from the origin of transfer in the region transferred early during bacterial conjugation. The cloned gene was able to suppress the UV and temperature sensitivity of an ssb-1 strain of Escherichia coli K-12. The nucleotide sequence of the ColIb ssb gene was determined, giving a predicted molecular weight of 19,110 for the SSB protein. Sequence data show that ColIb ssb is very similar to the ssb gene on plasmid F, which is also known to map in the leader region. High-level expression of ssb on ColIb required derepression of the transfer (tra) genes and the activity of the positive regulatory system controlling these genes, suggesting that the SSB protein contributes to the conjugative processing of DNA. A mutant of ColIbdrd-1 carrying a Tn903-derived insertion in ssb was constructed, but it was unaffected in the ability to generate plasmid transconjugants and it was maintained apparently stably in donor cells both following mating and during vegetative growth. Hence, no biological role of ColIb SSB protein was detected. However, unlike the parental plasmid, such ColIb ssb mutants conferred a marked Psi+ (plasmid-mediated SOS inhibition) phenotype on recA441 and recA730 strains, implying a functional relationship between SSB and Psi proteins.

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Selected References

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  1. Bagdasarian M., Bailone A., Bagdasarian M. M., Manning P. A., Lurz R., Timmis K. N., Devoret R. An inhibitor of SOS induction, specified by a plasmid locus in Escherichia coli. Proc Natl Acad Sci U S A. 1986 Aug;83(15):5723–5726. doi: 10.1073/pnas.83.15.5723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bagdasarian M., D'Ari R., Filipowicz W., George J. Suppression of induction of SOS functions in an Escherichia coli tif-1 mutant by plasmid R100.1. J Bacteriol. 1980 Feb;141(2):464–469. doi: 10.1128/jb.141.2.464-469.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bailone A., Bäckman A., Sommer S., Célérier J., Bagdasarian M. M., Bagdasarian M., Devoret R. PsiB polypeptide prevents activation of RecA protein in Escherichia coli. Mol Gen Genet. 1988 Nov;214(3):389–395. doi: 10.1007/BF00330471. [DOI] [PubMed] [Google Scholar]
  4. Boulnois G. J., Beddoes M. J., Wilkins B. M. Rifampin disrupts conjugal and chromosomal deoxyribonucleic acid metabolism in Escherichia coli K-12 carrying some IncIalpha plasmids. J Bacteriol. 1979 May;138(2):324–332. doi: 10.1128/jb.138.2.324-332.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bradley D. E. Morphological and serological relationships of conjugative pili. Plasmid. 1980 Sep;4(2):155–169. doi: 10.1016/0147-619x(80)90005-0. [DOI] [PubMed] [Google Scholar]
  6. Casaregola S., D'Ari R., Huisman O. Quantitative evaluation of recA gene expression in Escherichia coli. Mol Gen Genet. 1982;185(3):430–439. doi: 10.1007/BF00334135. [DOI] [PubMed] [Google Scholar]
  7. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chase J. W., Merrill B. M., Williams K. R. F sex factor encodes a single-stranded DNA binding protein (SSB) with extensive sequence homology to Escherichia coli SSB. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5480–5484. doi: 10.1073/pnas.80.18.5480. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chase J. W., Williams K. R. Single-stranded DNA binding proteins required for DNA replication. Annu Rev Biochem. 1986;55:103–136. doi: 10.1146/annurev.bi.55.070186.000535. [DOI] [PubMed] [Google Scholar]
  10. Chatfield L. K., Orr E., Boulnois G. J., Wilkins B. M. DNA primase of plasmid ColIb is involved in conjugal DnA synthesis in donor and recipient bacteria. J Bacteriol. 1982 Dec;152(3):1188–1195. doi: 10.1128/jb.152.3.1188-1195.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Chou P. Y., Fasman G. D. Prediction of the secondary structure of proteins from their amino acid sequence. Adv Enzymol Relat Areas Mol Biol. 1978;47:45–148. doi: 10.1002/9780470122921.ch2. [DOI] [PubMed] [Google Scholar]
  12. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Falkow S., Guerry P., Hedges R. W., Datta N. Polynucleotide sequence relationships among plasmids of the I compatibility complex. J Gen Microbiol. 1974 Nov;85(1):65–76. doi: 10.1099/00221287-85-1-65. [DOI] [PubMed] [Google Scholar]
  14. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  15. Gerdes K., Rasmussen P. B., Molin S. Unique type of plasmid maintenance function: postsegregational killing of plasmid-free cells. Proc Natl Acad Sci U S A. 1986 May;83(10):3116–3120. doi: 10.1073/pnas.83.10.3116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Glassberg J., Meyer R. R., Kornberg A. Mutant single-strand binding protein of Escherichia coli: genetic and physiological characterization. J Bacteriol. 1979 Oct;140(1):14–19. doi: 10.1128/jb.140.1.14-19.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Golub E. I., Low K. B. Conjugative plasmids of enteric bacteria from many different incompatibility groups have similar genes for single-stranded DNA-binding proteins. J Bacteriol. 1985 Apr;162(1):235–241. doi: 10.1128/jb.162.1.235-241.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Golub E. I., Low K. B. Derepression of single-stranded DNA-binding protein genes on plasmids derepressed for conjugation, and complementation of an E. coli ssb- mutation by these genes. Mol Gen Genet. 1986 Sep;204(3):410–416. doi: 10.1007/BF00331017. [DOI] [PubMed] [Google Scholar]
  19. Golub E., Bailone A., Devoret R. A gene encoding an SOS inhibitor is present in different conjugative plasmids. J Bacteriol. 1988 Sep;170(9):4392–4394. doi: 10.1128/jb.170.9.4392-4394.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Grindley N. D., Humphreys G. O., Anderson E. S. Molecular studies of R factor compatibility groups. J Bacteriol. 1973 Jul;115(1):387–398. doi: 10.1128/jb.115.1.387-398.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Holmes D. S., Quigley M. A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem. 1981 Jun;114(1):193–197. doi: 10.1016/0003-2697(81)90473-5. [DOI] [PubMed] [Google Scholar]
  22. Howland C. J., Wilkins B. M. Direction of conjugative transfer of IncI1 plasmid ColIb-P9. J Bacteriol. 1988 Oct;170(10):4958–4959. doi: 10.1128/jb.170.10.4958-4959.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Huisman O., D'Ari R. An inducible DNA replication-cell division coupling mechanism in E. coli. Nature. 1981 Apr 30;290(5809):797–799. doi: 10.1038/290797a0. [DOI] [PubMed] [Google Scholar]
  24. Huisman O., D'Ari R. Effect of suppressors of SOS-mediated filamentation on sfiA operon expression in Escherichia coli. J Bacteriol. 1983 Jan;153(1):169–175. doi: 10.1128/jb.153.1.169-175.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kolodkin A. L., Capage M. A., Golub E. I., Low K. B. F sex factor of Escherichia coli K-12 codes for a single-stranded DNA binding protein. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4422–4426. doi: 10.1073/pnas.80.14.4422. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  27. Lieberman H. B., Witkin E. M. DNA degradation, UV sensitivity and SOS-mediated mutagenesis in strains of Escherichia coli deficient in single-strand DNA binding protein: effects of mutations and treatments that alter levels of Exonuclease V or recA protein. Mol Gen Genet. 1983;190(1):92–100. doi: 10.1007/BF00330329. [DOI] [PubMed] [Google Scholar]
  28. Merryweather A., Rees C. E., Smith N. M., Wilkins B. M. Role of sog polypeptides specified by plasmid ColIb-P9 and their transfer between conjugating bacteria. EMBO J. 1986 Nov;5(11):3007–3012. doi: 10.1002/j.1460-2075.1986.tb04599.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Meyer R. R., Glassberg J., Kornberg A. An Escherichia coli mutant defective in single-strand binding protein is defective in DNA replication. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1702–1705. doi: 10.1073/pnas.76.4.1702. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Ogura T., Hiraga S. Mini-F plasmid genes that couple host cell division to plasmid proliferation. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4784–4788. doi: 10.1073/pnas.80.15.4784. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Oka A., Sugisaki H., Takanami M. Nucleotide sequence of the kanamycin resistance transposon Tn903. J Mol Biol. 1981 Apr 5;147(2):217–226. doi: 10.1016/0022-2836(81)90438-1. [DOI] [PubMed] [Google Scholar]
  32. Reed K. C., Mann D. A. Rapid transfer of DNA from agarose gels to nylon membranes. Nucleic Acids Res. 1985 Oct 25;13(20):7207–7221. doi: 10.1093/nar/13.20.7207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Rees C. E., Bradley D. E., Wilkins B. M. Organization and regulation of the conjugation genes of IncI1 plasmid colIb-P9. Plasmid. 1987 Nov;18(3):223–236. doi: 10.1016/0147-619x(87)90065-5. [DOI] [PubMed] [Google Scholar]
  34. Sancar A., Williams K. R., Chase J. W., Rupp W. D. Sequences of the ssb gene and protein. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4274–4278. doi: 10.1073/pnas.78.7.4274. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Soberon X., Covarrubias L., Bolivar F. Construction and characterization of new cloning vehicles. IV. Deletion derivatives of pBR322 and pBR325. Gene. 1980 May;9(3-4):287–305. doi: 10.1016/0378-1119(90)90328-o. [DOI] [PubMed] [Google Scholar]
  36. Stoker N. G., Fairweather N. F., Spratt B. G. Versatile low-copy-number plasmid vectors for cloning in Escherichia coli. Gene. 1982 Jun;18(3):335–341. doi: 10.1016/0378-1119(82)90172-x. [DOI] [PubMed] [Google Scholar]
  37. Taylor L. A., Rose R. E. A correction in the nucleotide sequence of the Tn903 kanamycin resistance determinant in pUC4K. Nucleic Acids Res. 1988 Jan 11;16(1):358–358. doi: 10.1093/nar/16.1.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  39. Way J. C., Davis M. A., Morisato D., Roberts D. E., Kleckner N. New Tn10 derivatives for transposon mutagenesis and for construction of lacZ operon fusions by transposition. Gene. 1984 Dec;32(3):369–379. doi: 10.1016/0378-1119(84)90012-x. [DOI] [PubMed] [Google Scholar]
  40. Whittier R. F., Chase J. W. DNA repair properties of Escherichia coli tif-1, recAo281 and lexA1 strains deficient in single-strand DNA binding protein. Mol Gen Genet. 1983;190(1):101–111. doi: 10.1007/BF00330330. [DOI] [PubMed] [Google Scholar]
  41. Wilkins B. M., Boulnois G. J., Lanka E. A plasmid DNA primase active in discontinuous bacterial DNA replication. Nature. 1981 Mar 19;290(5803):217–221. doi: 10.1038/290217a0. [DOI] [PubMed] [Google Scholar]
  42. Willetts N., Wilkins B. Processing of plasmid DNA during bacterial conjugation. Microbiol Rev. 1984 Mar;48(1):24–41. doi: 10.1128/mr.48.1.24-41.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Williams K. R., Spicer E. K., LoPresti M. B., Guggenheimer R. A., Chase J. W. Limited proteolysis studies on the Escherichia coli single-stranded DNA binding protein. Evidence for a functionally homologous domain in both the Escherichia coli and T4 DNA binding proteins. J Biol Chem. 1983 Mar 10;258(5):3346–3355. [PubMed] [Google Scholar]
  44. Winans S. C., Elledge S. J., Krueger J. H., Walker G. C. Site-directed insertion and deletion mutagenesis with cloned fragments in Escherichia coli. J Bacteriol. 1985 Mar;161(3):1219–1221. doi: 10.1128/jb.161.3.1219-1221.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]

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