Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1994 Jul;176(14):4316–4320. doi: 10.1128/jb.176.14.4316-4320.1994

Isolation and biochemical characterization of a novel lantibiotic mutacin from Streptococcus mutans.

J Novák 1, P W Caufield 1, E J Miller 1
PMCID: PMC205644  PMID: 8021218

Abstract

Certain members of the indigenous biota of humans produce antimicrobial substances called bacteriocins, which inhibit other bacteria, including members of their own species. One of these substances, mutacin, is made by Streptococcus mutans, a member of the oral biota. Mutacin inhibits other mutans streptococci as well as many gram-positive exogenous pathogens. Here, we report for the first time the purification and partial biochemical characterization of a lanthionine-containing mutacin peptide from S. mutants T8. The biologically active peptide was isolated from the broth cultures by ultrafiltration and differential precipitation. The final mutacin preparation was homogeneous as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and N-terminal amino acid sequencing. A molecular mass of the peptide was estimated by electrospray ionization mass spectroscopy to be 3,244.64 +/- 1.15 Da. Its amino acid composition indicates the presence of lanthionine and likely beta-methyllanthionine in a total of about 25 amino acids. Because alpha,beta-unsaturated amino acids, the precursors of lanthionine residues, are often found in lantibiotics, we carried out the addition reaction of the mutacin with N-(methyl)mercaptoacetamide. The subsequent electrospray ionization mass spectroscopy analysis indicated the presence of two reaction products with M(r)s of 3,350.45 and 3,456.0. These are interpreted as the mutacin molecule with the addition of one and two molecules of reagent to the unsaturated amino acids, respectively. Sequencing of the peptide revealed an N-terminal amino acid sequence of Asn-Arg-Trp-Trp-Gln-Gly-Val-Val.

Full text

PDF
4316

Images in this article

4318Image
on p.4318

Selected References

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

  1. Allgaier H., Jung G., Werner R. G., Schneider U., Zähner H. Epidermin: sequencing of a heterodetic tetracyclic 21-peptide amide antibiotic. Eur J Biochem. 1986 Oct 1;160(1):9–22. doi: 10.1111/j.1432-1033.1986.tb09933.x. [DOI] [PubMed] [Google Scholar]
  2. Berkowitz R. J., Jones P. Mouth-to-mouth transmission of the bacterium Streptococcus mutans between mother and child. Arch Oral Biol. 1985;30(4):377–379. doi: 10.1016/0003-9969(85)90014-7. [DOI] [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Buchman G. W., Banerjee S., Hansen J. N. Structure, expression, and evolution of a gene encoding the precursor of nisin, a small protein antibiotic. J Biol Chem. 1988 Nov 5;263(31):16260–16266. [PubMed] [Google Scholar]
  5. Caufield P. W., Childers N. K., Allen D. N., Hansen J. B. Distinct bacteriocin groups correlate with different groups of Streptococcus mutans plasmids. Infect Immun. 1985 Apr;48(1):51–56. doi: 10.1128/iai.48.1.51-56.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Caufield P. W., Shah G. R., Hollingshead S. K. Use of transposon Tn916 to inactivate and isolate a mutacin-associated gene from Streptococcus mutans. Infect Immun. 1990 Dec;58(12):4126–4135. doi: 10.1128/iai.58.12.4126-4135.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Caufield P. W., Shah G., Hollingshead S. K., Parrot M., Lavoie M. C. Evidence that mutacin II production is not mediated by a 5.6-kb plasmid in Streptococcus mutans. Plasmid. 1990 Sep;24(2):110–118. doi: 10.1016/0147-619x(90)90013-3. [DOI] [PubMed] [Google Scholar]
  8. Davey A. L., Rogers A. H. Multiple types of the bacterium Streptococcus mutans in the human mouth and their intra-family transmission. Arch Oral Biol. 1984;29(6):453–460. doi: 10.1016/0003-9969(84)90026-8. [DOI] [PubMed] [Google Scholar]
  9. Delisle A. L. Properties of mutacin b, an antibacterial substance produced by Streptococcus mutans strain BHT. Microbios. 1986;46(186):21–28. [PubMed] [Google Scholar]
  10. Gross E., Kiltz H. H. The number and nature of , -unsaturated amino acids in subtilin. Biochem Biophys Res Commun. 1973 Jan 23;50(2):559–565. doi: 10.1016/0006-291x(73)90876-0. [DOI] [PubMed] [Google Scholar]
  11. Gross E., Morell J. L. The number and nature of alpha,beta-unsaturated amino acids in nisin. FEBS Lett. 1968 Nov;2(1):61–64. doi: 10.1016/0014-5793(68)80101-2. [DOI] [PubMed] [Google Scholar]
  12. Gross E., Morell J. L. The structure of nisin. J Am Chem Soc. 1971 Sep 8;93(18):4634–4635. doi: 10.1021/ja00747a073. [DOI] [PubMed] [Google Scholar]
  13. Hamada S., Imanishi H., Ooshima T. Isolation and mode of action of a cell-free bacteriocin (mutacin) from serotype g Streptococcus mutans MT3791. Zentralbl Bakteriol Mikrobiol Hyg A. 1986 May;261(3):287–298. doi: 10.1016/s0176-6724(86)80046-3. [DOI] [PubMed] [Google Scholar]
  14. Hamada S., Ooshima T. Inhibitory spectrum of a bacteriocinlike substance (mutacin) produced by some strains of Streptococcus mutans. J Dent Res. 1975 Jan-Feb;54(1):140–145. doi: 10.1177/00220345750540010801. [DOI] [PubMed] [Google Scholar]
  15. Hamada S., Slade H. D. Biology, immunology, and cariogenicity of Streptococcus mutans. Microbiol Rev. 1980 Jun;44(2):331–384. doi: 10.1128/mr.44.2.331-384.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hillman J. D., Johnson K. P., Yaphe B. I. Isolation of a Streptococcus mutans strain producing a novel bacteriocin. Infect Immun. 1984 Apr;44(1):141–144. doi: 10.1128/iai.44.1.141-144.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Ikeda T., Iwanami T., Hirasawa M., Watanabe C., McGhee J. R., Shiota T. Purification and certain properties of a bacteriocin from Streptococcus mutans. Infect Immun. 1982 Mar;35(3):861–868. doi: 10.1128/iai.35.3.861-868.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kaletta C., Entian K. D., Kellner R., Jung G., Reis M., Sahl H. G. Pep5, a new lantibiotic: structural gene isolation and prepeptide sequence. Arch Microbiol. 1989;152(1):16–19. doi: 10.1007/BF00447005. [DOI] [PubMed] [Google Scholar]
  19. Kaletta C., Entian K. D. Nisin, a peptide antibiotic: cloning and sequencing of the nisA gene and posttranslational processing of its peptide product. J Bacteriol. 1989 Mar;171(3):1597–1601. doi: 10.1128/jb.171.3.1597-1601.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kelstrup J., Gibbons R. J. Bacteriocins from human and rodent streptococci. Arch Oral Biol. 1969 Mar;14(3):251–258. doi: 10.1016/0003-9969(69)90227-1. [DOI] [PubMed] [Google Scholar]
  21. Kurita T., Hirasawa M. Biological and biochemical characterization of novel lipid-like antibacterial substances (mutalipocins) produced by Streptoccus mutans strain 32K. J Gen Microbiol. 1988 Jan;134(1):213–220. doi: 10.1099/00221287-134-1-213. [DOI] [PubMed] [Google Scholar]
  22. Loyola-Rodriguez J. P., Morisaki I., Kitamura K., Hamada S. Purification and properties of extracellular mutacin, a bacteriocin from Streptococcus sobrinus. J Gen Microbiol. 1992 Feb;138(2):269–274. doi: 10.1099/00221287-138-2-269. [DOI] [PubMed] [Google Scholar]
  23. Miller E. J., Narkates A. J., Niemann M. A. Amino acid analysis of collagen hydrolysates by reverse-phase high-performance liquid chromatography of 9-fluorenylmethyl chloroformate derivatives. Anal Biochem. 1990 Oct;190(1):92–97. doi: 10.1016/0003-2697(90)90139-z. [DOI] [PubMed] [Google Scholar]
  24. Parrot M., Caufield P. W., Lavoie M. C. Preliminary characterization of four bacteriocins from Streptococcus mutans. Can J Microbiol. 1990 Feb;36(2):123–130. doi: 10.1139/m90-022. [DOI] [PubMed] [Google Scholar]
  25. Paul D., Slade H. D. Production and properties of an extracellular bacteriocin from Streptococcus mutans bacteriocidal for group A and other streptococci. Infect Immun. 1975 Dec;12(6):1375–1385. doi: 10.1128/iai.12.6.1375-1385.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Rogers A. H. Bacteriocin patterns of strains belonging to various serotypes of Streptococcus mutans. Arch Oral Biol. 1976;21(4):243–249. doi: 10.1016/0003-9969(76)90042-x. [DOI] [PubMed] [Google Scholar]
  27. Rogers A. H. Effect of the medium on bacteriocin production among strains of Streptococcus mutans. Appl Microbiol. 1972 Aug;24(2):294–295. doi: 10.1128/am.24.2.294-295.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Schnell N., Entian K. D., Schneider U., Götz F., Zähner H., Kellner R., Jung G. Prepeptide sequence of epidermin, a ribosomally synthesized antibiotic with four sulphide-rings. Nature. 1988 May 19;333(6170):276–278. doi: 10.1038/333276a0. [DOI] [PubMed] [Google Scholar]
  29. Schägger H., von Jagow G. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. Anal Biochem. 1987 Nov 1;166(2):368–379. doi: 10.1016/0003-2697(87)90587-2. [DOI] [PubMed] [Google Scholar]
  30. Tagg J. R., Ragland N. L., Dickson N. P. A longitudinal study of Lancefield group A streptococcus acquisitions by a group of young Dunedin schoolchildren. N Z Med J. 1990 Sep 12;103(897):429–431. [PubMed] [Google Scholar]
  31. Tagg J. R. Studies of "BLIS-ful" oral bacteria. N Z Dent J. 1991 Jan;87(387):14–16. [PubMed] [Google Scholar]
  32. van de Rijn I., Kessler R. E. Growth characteristics of group A streptococci in a new chemically defined medium. Infect Immun. 1980 Feb;27(2):444–448. doi: 10.1128/iai.27.2.444-448.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. van der Meer J. R., Polman J., Beerthuyzen M. M., Siezen R. J., Kuipers O. P., De Vos W. M. Characterization of the Lactococcus lactis nisin A operon genes nisP, encoding a subtilisin-like serine protease involved in precursor processing, and nisR, encoding a regulatory protein involved in nisin biosynthesis. J Bacteriol. 1993 May;175(9):2578–2588. doi: 10.1128/jb.175.9.2578-2588.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES