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Infection and Immunity logoLink to Infection and Immunity
. 1996 Oct;64(10):4137–4142. doi: 10.1128/iai.64.10.4137-4142.1996

Insertional inactivation of an intrageneric coaggregation-relevant adhesin locus from Streptococcus gordonii DL1 (Challis).

C J Whittaker 1, D L Clemans 1, P E Kolenbrander 1
PMCID: PMC174348  PMID: 8926080

Abstract

Transposon Tn916 was used to insertionally inactivate a coaggregation-relevant locus of Streptococcus gordonii DL1 (Challis). One mutant (F11) was isolated that lost the ability to coaggregate with the streptococcal partners of DL1 but retained the ability to coaggregate with partners belonging to other genera. A probe specific for the region flanking the Tn916 insertion was used to isolate a locus-specific fragment from a chromosomal lambda library. Southern analysis of the resulting phagemids revealed that a 0.5-kb EcoRI fragment hybridized with the F11 probe. Cloning of the 0.5-kb EcoRI fragment into the E. coli-streptococcal insertion vector p(omega) yielded pCW4, which was used to insertionally inactivate the putative coaggregation-relevant gene in DL1. Insertion mutants showed altered coaggregation with streptococci but retained wild-type coaggregation properties with other genera of bacteria. Comparison of immunoblots of cell surface proteins showed a 100-kDa protein in DL1 which was not detected in the Tn916 and pCW4 insertion mutants. These results indicate that the 0.5-kb EcoRI fragment is part of an adhesin-relevant locus that is involved in the production of a 100-kDa protein at the cell surface.

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Selected References

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  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  2. Andersen R. N., Ganeshkumar N., Kolenbrander P. E. Cloning of the Streptococcus gordonii PK488 gene, encoding an adhesin which mediates coaggregation with Actinomyces naeslundii PK606. Infect Immun. 1993 Mar;61(3):981–987. doi: 10.1128/iai.61.3.981-987.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baxter-Gabbard K. L. A simple method for the large-scale preparation of sucrose gradients. FEBS Lett. 1972 Jan 15;20(1):117–119. doi: 10.1016/0014-5793(72)80031-0. [DOI] [PubMed] [Google Scholar]
  4. Behnke D. Plasmid transformation of Streptococcus sanguis (Challis) occurs by circular and linear molecules. Mol Gen Genet. 1981;182(3):490–497. doi: 10.1007/BF00293940. [DOI] [PubMed] [Google Scholar]
  5. Chassy B. M. A gentle method for the lysis of oral streptococci. Biochem Biophys Res Commun. 1976 Jan 26;68(2):603–608. doi: 10.1016/0006-291x(76)91188-8. [DOI] [PubMed] [Google Scholar]
  6. Chassy B. M., Giuffrida A. Method for the lysis of Gram-positive, asporogenous bacteria with lysozyme. Appl Environ Microbiol. 1980 Jan;39(1):153–158. doi: 10.1128/aem.39.1.153-158.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Ciardi J. E., McCray G. F., Kolenbrander P. E., Lau A. Cell-to-cell interaction of Streptococcus sanguis and Propionibacterium acnes on saliva-coated hydroxyapatite. Infect Immun. 1987 Jun;55(6):1441–1446. doi: 10.1128/iai.55.6.1441-1446.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clemans D. L., Kolenbrander P. E. Identification of a 100-kilodalton putative coaggregation-mediating adhesin of Streptococcus gordonii DL1 (Challis). Infect Immun. 1995 Dec;63(12):4890–4893. doi: 10.1128/iai.63.12.4890-4893.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Clemans D. L., Kolenbrander P. E. Isolation and characterization of coaggregation-defective (Cog-) mutants of Streptococcus gordonii DL1 (Challis). J Ind Microbiol. 1995 Sep;15(3):193–197. doi: 10.1007/BF01569825. [DOI] [PubMed] [Google Scholar]
  10. Correia F. F., DiRienzo J. M., McKay T. L., Rosan B. scbA from Streptococcus crista CC5A: an atypical member of the lraI gene family. Infect Immun. 1996 Jun;64(6):2114–2121. doi: 10.1128/iai.64.6.2114-2121.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Demuth D. R., Duan Y., Brooks W., Holmes A. R., McNab R., Jenkinson H. F. Tandem genes encode cell-surface polypeptides SspA and SspB which mediate adhesion of the oral bacterium Streptococcus gordonii to human and bacterial receptors. Mol Microbiol. 1996 Apr;20(2):403–413. doi: 10.1111/j.1365-2958.1996.tb02627.x. [DOI] [PubMed] [Google Scholar]
  12. Fenno J. C., Shaikh A., Spatafora G., Fives-Taylor P. The fimA locus of Streptococcus parasanguis encodes an ATP-binding membrane transport system. Mol Microbiol. 1995 Mar;15(5):849–863. doi: 10.1111/j.1365-2958.1995.tb02355.x. [DOI] [PubMed] [Google Scholar]
  13. Fraser C. M., Gocayne J. D., White O., Adams M. D., Clayton R. A., Fleischmann R. D., Bult C. J., Kerlavage A. R., Sutton G., Kelley J. M. The minimal gene complement of Mycoplasma genitalium. Science. 1995 Oct 20;270(5235):397–403. doi: 10.1126/science.270.5235.397. [DOI] [PubMed] [Google Scholar]
  14. Ganeshkumar N., Arora N., Kolenbrander P. E. Saliva-binding protein (SsaB) from Streptococcus sanguis 12 is a lipoprotein. J Bacteriol. 1993 Jan;175(2):572–574. doi: 10.1128/jb.175.2.572-574.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ganeshkumar N., Hannam P. M., Kolenbrander P. E., McBride B. C. Nucleotide sequence of a gene coding for a saliva-binding protein (SsaB) from Streptococcus sanguis 12 and possible role of the protein in coaggregation with actinomyces. Infect Immun. 1991 Mar;59(3):1093–1099. doi: 10.1128/iai.59.3.1093-1099.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gawron-Burke C., Clewell D. B. Regeneration of insertionally inactivated streptococcal DNA fragments after excision of transposon Tn916 in Escherichia coli: strategy for targeting and cloning of genes from gram-positive bacteria. J Bacteriol. 1984 Jul;159(1):214–221. doi: 10.1128/jb.159.1.214-221.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jacob A. E., Horton W. A., Drucker D. B. Genetic transformation in some cariogenic Streptococcus milleri. Microbios. 1989;60(244-245):167–175. [PubMed] [Google Scholar]
  18. Jenkinson H. F., Baker R. A., Tannock G. W. A binding-lipoprotein-dependent oligopeptide transport system in Streptococcus gordonii essential for uptake of hexa- and heptapeptides. J Bacteriol. 1996 Jan;178(1):68–77. doi: 10.1128/jb.178.1.68-77.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jenkinson H. F. Cell surface protein receptors in oral streptococci. FEMS Microbiol Lett. 1994 Aug 15;121(2):133–140. doi: 10.1111/j.1574-6968.1994.tb07089.x. [DOI] [PubMed] [Google Scholar]
  20. Jenkinson H. F., Easingwood R. A. Insertional inactivation of the gene encoding a 76-kilodalton cell surface polypeptide in Streptococcus gordonii Challis has a pleiotropic effect on cell surface composition and properties. Infect Immun. 1990 Nov;58(11):3689–3697. doi: 10.1128/iai.58.11.3689-3697.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jenkinson H. F., Terry S. D., McNab R., Tannock G. W. Inactivation of the gene encoding surface protein SspA in Streptococcus gordonii DL1 affects cell interactions with human salivary agglutinin and oral actinomyces. Infect Immun. 1993 Aug;61(8):3199–3208. doi: 10.1128/iai.61.8.3199-3208.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kolenbrander P. E., Andersen R. N. Characterization of Streptococcus gordonii (S. sanguis) PK488 adhesin-mediated coaggregation with Actinomyces naeslundii PK606. Infect Immun. 1990 Sep;58(9):3064–3072. doi: 10.1128/iai.58.9.3064-3072.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kolenbrander P. E., Andersen R. N., Ganeshkumar N. Nucleotide sequence of the Streptococcus gordonii PK488 coaggregation adhesin gene, scaA, and ATP-binding cassette. Infect Immun. 1994 Oct;62(10):4469–4480. doi: 10.1128/iai.62.10.4469-4480.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kolenbrander P. E., Andersen R. N., Moore L. V. Intrageneric coaggregation among strains of human oral bacteria: potential role in primary colonization of the tooth surface. Appl Environ Microbiol. 1990 Dec;56(12):3890–3894. doi: 10.1128/aem.56.12.3890-3894.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kolenbrander P. E. Isolation and characterization of coaggregation-defective mutants of Actinomyces viscosus, Actinomyces naeslundii, and Streptococcus sanguis. Infect Immun. 1982 Sep;37(3):1200–1208. doi: 10.1128/iai.37.3.1200-1208.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kolenbrander P. E., London J. Adhere today, here tomorrow: oral bacterial adherence. J Bacteriol. 1993 Jun;175(11):3247–3252. doi: 10.1128/jb.175.11.3247-3252.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kolenbrander P. E., Parrish K. D., Andersen R. N., Greenberg E. P. Intergeneric coaggregation of oral Treponema spp. with Fusobacterium spp. and intrageneric coaggregation among Fusobacterium spp. Infect Immun. 1995 Dec;63(12):4584–4588. doi: 10.1128/iai.63.12.4584-4588.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Laemmli U. K., Favre M. Maturation of the head of bacteriophage T4. I. DNA packaging events. J Mol Biol. 1973 Nov 15;80(4):575–599. doi: 10.1016/0022-2836(73)90198-8. [DOI] [PubMed] [Google Scholar]
  29. Lamont R. J., Demuth D. R., Davis C. A., Malamud D., Rosan B. Salivary-agglutinin-mediated adherence of Streptococcus mutans to early plaque bacteria. Infect Immun. 1991 Oct;59(10):3446–3450. doi: 10.1128/iai.59.10.3446-3450.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lamont R. J., Gil S., Demuth D. R., Malamud D., Rosan B. Molecules of Streptococcus gordonii that bind to Porphyromonas gingivalis. Microbiology. 1994 Apr;140(Pt 4):867–872. doi: 10.1099/00221287-140-4-867. [DOI] [PubMed] [Google Scholar]
  31. Liljemark W. F., Bloomquist C. G., Coulter M. C., Fenner L. J., Skopek R. J., Schachtele C. F. Utilization of a continuous streptococcal surface to measure interbacterial adherence in vitro and in vivo. J Dent Res. 1988 Dec;67(12):1455–1460. doi: 10.1177/00220345880670120301. [DOI] [PubMed] [Google Scholar]
  32. Lowe A. M., Lambert P. A., Smith A. W. Cloning of an Enterococcus faecalis endocarditis antigen: homology with adhesins from some oral streptococci. Infect Immun. 1995 Feb;63(2):703–706. doi: 10.1128/iai.63.2.703-706.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Lunsford R. D. A Tn4001 delivery system for Streptococcus gordonii (Challis). Plasmid. 1995 Mar;33(2):153–157. doi: 10.1006/plas.1995.1016. [DOI] [PubMed] [Google Scholar]
  34. Mandel M., Higa A. Calcium-dependent bacteriophage DNA infection. J Mol Biol. 1970 Oct 14;53(1):159–162. doi: 10.1016/0022-2836(70)90051-3. [DOI] [PubMed] [Google Scholar]
  35. Maryanski J. H., Wittenberger C. L. Mannitol transport in Streptococcus mutans. J Bacteriol. 1975 Dec;124(3):1475–1481. doi: 10.1128/jb.124.3.1475-1481.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. McNab R., Jenkinson H. F. Gene disruption identifies a 290 kDa cell-surface polypeptide conferring hydrophobicity and coaggregation properties in Streptococcus gordonii. Mol Microbiol. 1992 Oct;6(20):2939–2949. doi: 10.1111/j.1365-2958.1992.tb01753.x. [DOI] [PubMed] [Google Scholar]
  37. Moriya S., Ogasawara N., Yoshikawa H. Structure and function of the region of the replication origin of the Bacillus subtilis chromosome. III. Nucleotide sequence of some 10,000 base pairs in the origin region. Nucleic Acids Res. 1985 Apr 11;13(7):2251–2265. doi: 10.1093/nar/13.7.2251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Morrison D. A., Trombe M. C., Hayden M. K., Waszak G. A., Chen J. D. Isolation of transformation-deficient Streptococcus pneumoniae mutants defective in control of competence, using insertion-duplication mutagenesis with the erythromycin resistance determinant of pAM beta 1. J Bacteriol. 1984 Sep;159(3):870–876. doi: 10.1128/jb.159.3.870-876.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Nyvad B., Kilian M. Microbiology of the early colonization of human enamel and root surfaces in vivo. Scand J Dent Res. 1987 Oct;95(5):369–380. doi: 10.1111/j.1600-0722.1987.tb01627.x. [DOI] [PubMed] [Google Scholar]
  40. Ogasawara N., Nakai S., Yoshikawa H. Systematic sequencing of the 180 kilobase region of the Bacillus subtilis chromosome containing the replication origin. DNA Res. 1994;1(1):1–14. doi: 10.1093/dnares/1.1.1. [DOI] [PubMed] [Google Scholar]
  41. Sampson J. S., O'Connor S. P., Stinson A. R., Tharpe J. A., Russell H. Cloning and nucleotide sequence analysis of psaA, the Streptococcus pneumoniae gene encoding a 37-kilodalton protein homologous to previously reported Streptococcus sp. adhesins. Infect Immun. 1994 Jan;62(1):319–324. doi: 10.1128/iai.62.1.319-324.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Skopek R. J., Liljemark W. F., Bloomquist C. G., Rudney J. D. Dental plaque development on defined streptococcal surfaces. Oral Microbiol Immunol. 1993 Feb;8(1):16–23. doi: 10.1111/j.1399-302x.1993.tb00537.x. [DOI] [PubMed] [Google Scholar]
  44. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]

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