1887

Abstract

The strongly hydrophobic bacteriocin amylovorin L471 from DCE 471 was isolated and purified to homogeneity from complex culture broth by a novel, rapid and simple three-step protocol including (i) ammonium sulphate precipitation, (ii) chloroform/methanol extraction/precipitation and (iii) reversed-phase HPLC, the only chromatographic step involved. The molecular mass of the peptide was determined to be 48769 Da by electrospray mass spectrometric analysis. N-terminal amino acid sequencing identified 35 amino acid residues as being identical to the N-terminal sequence of lactobin A, a bacteriocin from another strain. These non-identical strains produce bacteriocins that display small differences in molecular mass and inhibitory spectrum. The amino acid sequence of amylovorin L471 shared significant homology with lactacin X, one of the two bactericidal peptides produced by VPI11088. A purified amylovorin L471 preparation permitted confirmation of the inhibitory spectrum previously established with a crude extract. It displayed a bactericidal mode of action on lactobacilli after an extremely rapid adsorption to the target cells. Two spp. were only weakly sensitive. Amylovorin L471 appears to be produced constitutively. Ethanol not only stimulated specific bacteriocin production but also prevented adsorption of the bacteriocin molecules to the producer cells upon prolonged fermentation. The latter result supports the hypothesis that the apparent inactivation of bacteriocin observed during the stationary phase of batch fermentations is due to adsorption.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-145-9-2559
1999-09-01
2019-10-24
Loading full text...

Full text loading...

/deliver/fulltext/micro/145/9/1452559a.html?itemId=/content/journal/micro/10.1099/00221287-145-9-2559&mimeType=html&fmt=ahah

References

  1. Allison, G. E., Frémaux, C. & Klaenhammer, T. R. ( 1994; ). Expansion of bacteriocin activity and host range upon complementation of two peptides encoded within the lactacin F operon. J Bacteriol 176, 2235-2241.
    [Google Scholar]
  2. Anderssen, E. L., Diep, D. B., Nes, I. F., Eijsink, V. G. H. & Nissen-Meyer, J. ( 1998; ). Antagonistic activity of Lactobacillus plantarum C11: two new two-peptide bacteriocins, plantaricin EF and JK, and the induction factor plantaricin A. Appl Environ Microbiol 64, 2269-2272.
    [Google Scholar]
  3. Atrih, A., Rekhif, N., Milliere, J. B. & Lefebvre, G. ( 1993; ). Detection and characterization of a bacteriocin produced by Lactobacillus plantarum C19. Can J Microbiol 39, 1173-1179.[CrossRef]
    [Google Scholar]
  4. Barefoot, S. F., Chen, Y.-R., Hughes, T. A., Bodine, A. B., Shearer, M. Y. & Hughes, M. D. ( 1994; ). Identification and purification of a protein that induces production of the Lactobacillus acidophilus bacteriocin lactacin B. Appl Environ Microbiol 60, 3522-3528.
    [Google Scholar]
  5. Bhunia, A. K., Johnson, M. C., Ray, B. & Kalchayanand, N. ( 1991; ). Mode of action of pediocin AcH from Pediococcus acidilactici H on sensitive bacterial strains. J Appl Bacteriol 70, 25-30.[CrossRef]
    [Google Scholar]
  6. Casaus, P., Nilsen, T., Cintas, L. M., Nes, I. F., Hernández, P. E. & Holo, H. ( 1997; ). Enterocin B, a new bacteriocin from Enterococcus faecium T136 which can act synergistically with enterocin A. Microbiology 143, 2287-2294.[CrossRef]
    [Google Scholar]
  7. Contreras, B. G. L., De Vuyst, L., Devreese, B., Busanyova, K., Raymaeckers, J., Bosman, F., Sablon, E. & Vandamme, E. J. ( 1997; ). Isolation, purification, and amino acid sequence of lactobin A, one of the two bacteriocins produced by Lactobacillus amylovorus LMG P-13139. Appl Environ Microbiol 63, 13-20.
    [Google Scholar]
  8. Daba, H., Lacroix, C., Huang, R. E. & Lemieux, L. ( 1994; ). Simple method of purification and sequencing of a bacteriocin produced by Pediococcus acidilactici UL5. J Appl Bacteriol 77, 682-688.[CrossRef]
    [Google Scholar]
  9. De Vos, W. M., Kuipers, O. M., van der Meer, J. R. & Siezen, R. J. ( 1995; ). Maturation pathway of nisin and other lantibiotics: posttranslationally modified antimicrobial peptides exported by Gram-positive bacteria. Mol Microbiol 17, 427-437.[CrossRef]
    [Google Scholar]
  10. De Vuyst, L. & Callewaert, R. (1997). Purification and large-scale isolation of amylovorin L471 and other class II bacteriocins. 8ème Colloque du Club des Bactéries Lactiques, ENSBANA, Université de Bourgogne, Dijon, France, aff. B3.
  11. De Vuyst, L. & Vandamme, E. J. (1994). Bacteriocins of Lactic Acid Bacteria: Microbiology, Genetics and Applications. London: Blackie Academic & Professional.
  12. De Vuyst, L., Callewaert, R. & Pot, B. ( 1996a; ). Characterization of the antagonistic activity of Lactobacillus amylovorus DCE 471 and large scale isolation of its bacteriocin amylovorin L471. Syst Appl Microbiol 19, 9-20.[CrossRef]
    [Google Scholar]
  13. De Vuyst, L., Callewaert, R. & Crabbé, K. ( 1996b; ). Primary metabolite kinetics of bacteriocin biosynthesis by Lactobacillus amylovorus and evidence for stimulation of bacteriocin production under unfavourable growth conditions. Microbiology 142, 817-827.[CrossRef]
    [Google Scholar]
  14. Diep, D. B., Håvarstein, L. S. & Nes, I. F. ( 1995; ). A bacteriocin-like peptide induces bacteriocin synthesis in Lactobacillus plantarum C11. Mol Microbiol 18, 631-639.[CrossRef]
    [Google Scholar]
  15. Elegado, F. B., Kim, W. J. & Kwon, D. Y. ( 1997; ). Rapid purification, partial characterization, and antimicrobial spectrum of the bacteriocin, pediocin AcM, from Pediococcus acidilactici M. Int J Food Microbiol 37, 1-11.[CrossRef]
    [Google Scholar]
  16. Ennahar, S., Aoude-Werner, D., Sorokine, O., van Dorsselaer, A., Bringel, F., Hubert, J. C. & Hasselmann, C. ( 1996; ). Production of pediocin AcH by Lactobacillus plantarum WHE 92 isolated from cheese. Appl Environ Microbiol 62, 4381-4387.
    [Google Scholar]
  17. Felix, J. V., Papathanasopoulos, M. A., Smith, A. A., von Holy, A. & Hastings, J. W. ( 1994; ). Characterization of leucocin B-Ta11a: a bacteriocin from Leuconostoc carnosum Ta11a isolated from meat. Curr Microbiol 29, 207-212.[CrossRef]
    [Google Scholar]
  18. Fenn, J. B., Mann, M., Meng, C. K., Wong, S. F. & Whitehouse, C. M. ( 1990; ). Electrospray ionization – principles and practice. Mass Spectrom Rev 9, 37-70.[CrossRef]
    [Google Scholar]
  19. Frémaux, C., Ahn, C. & Klaenhammer, T. R. ( 1993; ). Molecular analysis of the lactacin F operon. Appl Environ Microbiol 59, 3906-3915.
    [Google Scholar]
  20. Gálvez, A., Valdivia, E., Abriouel, H., Camafeita, E., Mendez, E., Martı́nez-Bueno, M. & Maqueda, M. ( 1998; ). Isolation and characterisation of enterocin EJ97, a bacteriocin produced by Enterococcus faecalis EJ97. Arch Microbiol 171, 59-65.[CrossRef]
    [Google Scholar]
  21. Hastings, J. M., Sailer, M., Johnson, K. L., Ray, K. L., Vederas, J. C. & Stiles, M. E. ( 1991; ). Characterization of leucocin A-UAL 187 and cloning of the bacteriocin gene from Leuconostoc gelidum. J Bacteriol 173, 7491-7500.
    [Google Scholar]
  22. Holck, A., Axelsson, L., Birkeland, S.-E., Aukrust, T. & Blom, H. ( 1992; ). Purification and amino acid sequence of sakacin A, a bacteriocin from Lactobacillus sake Lb706. J Gen Microbiol 138, 2715-2720.[CrossRef]
    [Google Scholar]
  23. Joerger, M. C. & Klaenhammer, T. R. ( 1986; ). Characterization and purification of helveticin J and evidence for a chromosomally determined bacteriocin produced by Lactobacillus helveticus 481. J Bacteriol 167, 439-446.
    [Google Scholar]
  24. Joosten, H. M., Nunez, M., Devreese, B., Van Beeumen, J. & Marugg, J. D. ( 1996; ). Purification and characterization of enterocin 4, a bacteriocin produced by Enterococcus faecalis INIA 4. Appl Environ Microbiol 62, 4220-4223.
    [Google Scholar]
  25. Klaenhammer, T. R. ( 1988; ). Bacteriocins of lactic acid bacteria. Biochimie 70, 337-349.[CrossRef]
    [Google Scholar]
  26. Klaenhammer, T. R. ( 1993; ). Genetics of bacteriocins produced by lactic acid bacteria. FEMS Microbiol Rev 12, 39-85.[CrossRef]
    [Google Scholar]
  27. Kyte, J. & Doolittle, R. F. ( 1982; ). A simple method for displaying the hydropathic character of a protein. J Mol Biol 157, 105-132.[CrossRef]
    [Google Scholar]
  28. Manca de Nadra, M. C., Sendino de Lamelas, D. & Strasser de Saad, A. M. ( 1998; ). Pediocin N5p from Pediococcus pentosaceus: adsorption on bacterial strains. Int J Food Microbiol 39, 79-85.[CrossRef]
    [Google Scholar]
  29. Mørtvedt-Abildgaard, C. I., Nissen-Meyer, J., Jelle, B., Grenov, B., Skaugen, M. & Nes, I. F. ( 1995; ). Production and pH-dependent bactericidal activity of lactocin S, a lantibiotic from Lactobacillus sake L45. Appl Environ Microbiol 61, 175-179.
    [Google Scholar]
  30. Motlagh, A. M., Bhunia, A. K., Szostek, F., Hansen, T. R., Johnson, M. G. & Ray, B. ( 1992; ). Nucleotide and amino acid sequence of pap-gene (pediocin AcH production) in Pediococcus acidilactici H. Lett Appl Microbiol 15, 45-48.[CrossRef]
    [Google Scholar]
  31. Muriana, P. M. & Klaenhammer, T. R. ( 1987; ). Conjugal transfer of plasmid-encoded determinants for bacteriocin production and immunity in Lactobacillus acidophilus 88. Appl Environ Microbiol 53, 553-560.
    [Google Scholar]
  32. Muriana, P. M. & Klaenhammer, T. R. ( 1991; ). Purification and partial characterisation of lactacin F, a bacteriocin produced by Lactobacillus acidophilus 11088. Appl Environ Microbiol 57, 114-121.
    [Google Scholar]
  33. Nes, I. F., Diep, D. B., Håvarstein, I. S., Brurberg, M. B., Eijsink, V. & Holo, H. ( 1996; ). Biosynthesis of bacteriocins in lactic acid bacteria. Antonie Leeuwenhoek 70, 113-128.[CrossRef]
    [Google Scholar]
  34. Nieto-Lozano, J. C., Meyer, J. N., Sletten, K., Peláz, C. & Nes, I. F. ( 1992; ). Purification and amino acid sequence of a bacteriocin produced by Pediococcus acidilactici. J Gen Microbiol 138, 1985-1990.[CrossRef]
    [Google Scholar]
  35. Nilsen, T., Nes, I. F. & Holo, H. ( 1998; ). An exported inducer peptide regulates bacteriocin production in Enterococcus faecium CTC492. J Bacteriol 180, 1848-1854.
    [Google Scholar]
  36. Nissen-Meyer, J., Holo, H., Håvarstein, L. S., Sletten, K. N. & Nes, I. F. ( 1992; ). A novel lactococcal bacteriocin whose activity depends on the complementary action of two peptides. J Bacteriol 174, 5686-5692.
    [Google Scholar]
  37. Papathanasopoulos, M. A., Dykes, G. A., Revol-Junelles, A.-M., Delfour, A., von Holy, A. & Hastings, J. W. ( 1998; ). Sequence and structural relationships of leucocins A-, B- and C-TA33a from Leuconostoc mesenteroides TA33a. Microbiology 144, 1343-1348.[CrossRef]
    [Google Scholar]
  38. Rammelsberg, M., Muller, E. & Radler, F. ( 1990; ). Caseicin 80: purification and characterization of a new bacteriocin from Lactobacillus casei. Arch Microbiol 154, 249-252.[CrossRef]
    [Google Scholar]
  39. Rekhif, N., Atrih, A. & Lefebvre, G. ( 1994; ). Characterisation and partial purification of plantaricin LC74, a bacteriocin produced by Lactobacillus plantarum LC74. Biotechnol Lett 16, 771-776.[CrossRef]
    [Google Scholar]
  40. Saucier, L., Poon, A. & Stiles, M. E. ( 1995; ). Induction of bacteriocin in Carnobacterium piscicola LV17. J Appl Bacteriol 78, 684-690.[CrossRef]
    [Google Scholar]
  41. Stoffels, G., Sahl, H.-G. & Gudmundsdóttir, Á. ( 1993; ). Carnocin UI49, a potential biopreservative produced by Carnobacterium piscicola: large scale purification and activity against various Gram-positive bacteria including Listeria sp. Int J Food Microbiol 20, 199-210.[CrossRef]
    [Google Scholar]
  42. Thompson, J. D., Higgins, D. G. & Gibson, T. J. ( 1994; ). clustal w: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22, 4673-4680.[CrossRef]
    [Google Scholar]
  43. Thompson, J. K., Collins, M. A. & Mercer, W. D. ( 1996; ). Characterization of a proteinaceous antimicrobial produced by Lactobacillus helveticus CNRZ450. J Appl Bacteriol 80, 338-348.[CrossRef]
    [Google Scholar]
  44. Tichaczek, P. S., Nissen-Meyer, J., Nes, I. F., Vogel, R. F. & Hammes, W. P. ( 1992; ). Characterization of the bacteriocins curvacin A from Lactobacillus curvatus LTH1174 and sakacin P from Lactobacillus sake LTH673. Syst Appl Microbiol 15, 460-468.[CrossRef]
    [Google Scholar]
  45. Vaughan, E. E., Daly, C. & Fitzgerald, G. F. ( 1992; ). Identification and characterization of helveticin V-1829, a bacteriocin produced by Lactobacillus helveticus 1829. J Appl Bacteriol 73, 299-308.[CrossRef]
    [Google Scholar]
  46. Venema, K., Chikindas, M. L., Seegers, J. F. M. L., Haandrikman, A. J., Leenhouts, K. J., Venema, G. & Kok, J. ( 1997; ). Rapid and efficient purification method for small, hydrophobic, cationic bacteriocins: purification of lactococcin B and pediocin PA-1. Appl Environ Microbiol 63, 305-309.
    [Google Scholar]
  47. Worobo, R. W., van Belkum, M. J., Sailer, M., Roy, K. L., Vederas, J. C. & Stiles, M. E. ( 1995; ). A signal peptide secretion-dependent bacteriocin from Carnobacterium divergens. J Bacteriol 177, 3143-3149.
    [Google Scholar]
  48. Yang, R., Johnson, M. C. & Ray, B. ( 1992; ). Novel method to extract large amounts of bacteriocins from lactic acid bacteria. Appl Environ Microbiol 58, 3355-3359.
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-145-9-2559
Loading
/content/journal/micro/10.1099/00221287-145-9-2559
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error