1887

Abstract

Germination of spores of T and 168 was inhibited by the trypsin inhibitors leupeptin and tosyllysine chloromethyl ketone (TLCK) and by the substrates tosylarginine methyl ester (TAME), benzoyl--arginine--nitroanilide (L-BAPNA) and D-BAPNA. Potencies of these inhibitory compounds were estimated by finding the concentration which inhibited 50 % germination (ID), as measured by events occurring early (loss of heat resistance), at an intermediate stage [dipicolinic acid (DPA) release], and late in germination (decrease in optical density). In T, all the compounds inhibited early and late events with the same ID. In , TAME inhibited early and late events at the same ID, but all other inhibitors had a lower ID for late events than for early events. This suggests that a trypsin-like enzyme activity is involved at two sequential stages in the germination of spores, one occurring at or before the loss of heat resistance and one at or before the decrease in optical density. Different trypsin-like activities were detected in broken dormant spores and germinated spores of T and in germinated spores of by means of three chromogenic substrates: benzoyI--phenylalanyl--valyl--arginine--nitroaniIid (L-PheVA), -BAPNA and -BAPNA. Separation of extracts of germinated spores on non-denaturing polyacrylamide gels showed that in both species the substrates were hydrolysed by three distinct enzymes with different electrophoretic mobilities. The three enzymes had different values for the above inhibitors. The possibility that -BAPNase from T might be involved in the initial germination steps was suggested by the similarity of ID values for germination and values for inhibition of the enzyme by TAME and TLCK, and by the fact that both germination and the -BAPNase were reversibly inhibited by TLCK.

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1991-05-01
2021-10-23
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References

  1. Boschwitz H. , Milner Y. , Keynan A. , Halvorson H. O. , Troll W. . 1983; Effect of inhibitors of trypsin-like proteolytic enzymes on Bacillus cereus T spore germination. Journal of Bacteriology 153 700 708
    [Google Scholar]
  2. Boschwitz H. , Halvorson H. O. , Kevnan A , Milner Y . 1985; Trypsinlike enzymes from dormant and germinated spores of Bacillus cereus T and their possible involvement in germination. Journal of Bacteriology 164 302 309
    [Google Scholar]
  3. Foster S. J. , Johnstone K. . 1986; The use of inhibitors to identify early events during Bacillus megaterium KM spore germination. Biochemical Journal 237 865 870
    [Google Scholar]
  4. Foster S. J. , Johnstone K. . 1987; Purification and properties of a germination-specific cortex-lytic enzyme from spores of Bacillus megaterium KM. Biochemical Journal 242 573 579
    [Google Scholar]
  5. Foster S. J. , Johnstone K. . 1988; Germination-specific cortexlytic enzyme is activated during triggering of Bacillus megaterium KM spore germination. Molecular Microbiology 2 727 733
    [Google Scholar]
  6. Gofshtein-Gandman L. V. , Keynan A. , Milner Y . 1988; Bacteria of the genus Bacillus have a hydrolase stereospecific to the D isomer of benzoyl-arginine-pnitroanilide. Journal of Bacteriology 170 5895 5900
    [Google Scholar]
  7. Gould G. W. , Hitchins A. D. . 1965; Germination of spores with Strange and Dark’s spore lytic enzyme. Spores III, 213 221 L. L. Campbell , H. O. Halvorson , Ann Arbor . Michigan: American Society for Microbiology;
    [Google Scholar]
  8. Gould G. W. , King W. L. . 1969; Action and properties of spore germination enzymes. Spores IV 276 286 L. L. Campbell , Bethesda . Maryland: American Society for Microbiology;
    [Google Scholar]
  9. Janoff A. S. , Coughlin R. T. , Racine F. M. , McGroarty E. J. , Vary J. C. . 1979; Use of electron spin resonance to study Bacillus megaterium spore membranes. Biochemical and Biophysical Research Communications 89 565 570
    [Google Scholar]
  10. Janssen F. W. , Lund A. J. , Anderson L. E. . 1958; Colorimetric assay for dipicolinic acid in bacterial spores. Science 127 26 27
    [Google Scholar]
  11. Keynan A. . 1978; Spore structure and its relations to resistance, dormancy, and germination. Spores VII, 43 53 G. Chambliss , J. C. Vary . Washington, DC; American Society for Microbiology:
    [Google Scholar]
  12. Keynan A. , Evenchick Z. , Halvorson H. O. , Hastings J. W. . 1964; Activation of bacterial endospores. Journal of Bacteriology 88 313 318
    [Google Scholar]
  13. Moir A. , Smith D. A. . 1985; The genetics of spore germination in Bacillus subtilis . Fundamental and Applied Aspects of Bacterial Spores 89 100 G. J. Dring , D. J. Ellar , G. W. Gould . London: Academic Press;
    [Google Scholar]
  14. Moir A. , Lafferty E. , Smith D. A. . 1979; Genetic analysis of spore germination mutants of Bacillus subtilis 168: the correlation of phenotype with map location. Journal of General Microbiology 111 165 180
    [Google Scholar]
  15. Piggot P. J. , Moir A. , Smith D. A. . 1981; Advances in the genetics of Bacillus subtilis differentiation. Sporulation and Germination, 29 39 H. S. Levinson , A. L. Sonenshein , D. J. Tipper . Washington, DC: American Society for Microbiology;
    [Google Scholar]
  16. Reusch V. M. , Hale S. G. , Hurly B. J. . 1982; Levels of cell wall enzymes in endospores and vegetative cells of Bacillus subtilis . Journal of Bacteriology 152 1147 1153
    [Google Scholar]
  17. Rossignol D. P. , Vary J. C. . 1977; A unique method for studying the initiation of Bacillus megaterium spore germination. Biochemical and Biophysical Research Communications 79 1098 1103
    [Google Scholar]
  18. Scott I. R. , Ellar D. J. . 1978; Study of calcium dipicolinate release during bacterial spore germination by using a new, sensitive assay for dipicolinate. Journal of Bacteriology 135 133 137
    [Google Scholar]
  19. Setlow P. . 1981; Biochemistry of bacterial forespore development and spore germination. Sporulation and Germination 13 28 H. S. Levinson , A. L. Sonenshein , D. J. Tipper . Washington, DC: American Society for Microbiology;
    [Google Scholar]
  20. Shaw E. , Mares-Guia M. , Cohen W. . 1965; Evidence for an active-center histidine in trypsin through use of specific reagent, 1-chloro-3-tosylamido-7-amino-2-heptanone, the chloromethyl ketone derived from N°-tosyl-llysine. Biochemistry 4 2219 2224
    [Google Scholar]
  21. Srivastava O. P. , Fitz-James P. . 1981; Isolation and characterization of a unique serine protease from Bacillus cereus spore coats. Journal of Gereral Microbiology 126 463 475
    [Google Scholar]
  22. Sterlini J. M. , Mandelstam J. . 1969; Commitment to sporulation in Bacillus subtilis and its relationship to development of actinomycin resistance. Biochemical Journal 113 29 37
    [Google Scholar]
  23. Stewart G. S. A. B. , Johnstone K. , Hagelberg E. , Ellar D. J. . 1981; Commitment of bacterial spores to germinate. Biochemical Journal 198 101 106
    [Google Scholar]
  24. Vary J. C. . 1978; Glucose-initiated germination in Bacillus megaterium spores. Spores VII, 104 108 G. Chambliss , J. C. Vary . Washington, DC: American Society for Microbiology;
    [Google Scholar]
  25. Vankatasubramanian P. , Johnstone K. . 1989; Biochemical analysis of the Bacillus subtilis 1604 spore germination response. Journal of General Microbiology 135 2723 2733
    [Google Scholar]
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