Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1985 Oct;164(1):302–309. doi: 10.1128/jb.164.1.302-309.1985

Trypsinlike enzymes from dormant and germinated spores of Bacillus cereus T and their possible involvement in germination.

H Boschwitz, H O Halvorson, A Keynan, Y Milner
PMCID: PMC214244  PMID: 3930468

Abstract

Trypsin-like enzymes were studied in dormant, activated, and germinated spores of Bacillus cereus T. Dormant spores contained two heat-labile enzyme activities. One was extractable with 2 M KCl and hydrolyzed azo-albumin. The second, a trypsinlike activity, was not extractable with 2 M KCl and hydrolyzed benzoyl-L-arginine-p-nitroanilide. Because of their heat instability, these two enzyme activities are probably not involved in the germination of heat-activated spores. Upon germination of heat-treated spores, a trypsinlike protease which was not detected in intact dormant spores was activated or exposed. This enzyme, when measured in intact germinated spores, hydrolyzed benzoyl-DL-arginine-p-nitroanilide but not azo-albumin and was inhibited in situ by sulfhydryl-blocking reagents such as p-chloromercuribenzoic acid and Hg2+. There was a correlation between the inhibition of germination and enzymatic activity by sulfhydryl-blocking reagents. The enzyme was also inhibited by leupeptin, tosyl-L-lysine chromoethyl ketone, and tosyl-L-arginine methyl ester. Good correlation existed between the inhibition of germination and enzymatic activity by these agents. Electron micrographs showed that in the presence of trypsin inhibitors, the spores did not lose their cortex. The protein extracts of the inhibited spores formed a somewhat different electrophoretic pattern in sodium dodecyl sulfate-polyacrylamide gel electrophoresis than the protein extracts of dormant or germinated spores.

Full text

PDF
302

Images in this article

307Image
on p.307
308Image
on p.308

Selected References

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

  1. Aronson A. I., Fitz-James P. C. Properties of Bacillus cereus spore coat mutants. J Bacteriol. 1975 Jul;123(1):354–365. doi: 10.1128/jb.123.1.354-365.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aronson A. I., Fitz-James P. Structure and morphogenesis of the bacterial spore coat. Bacteriol Rev. 1976 Jun;40(2):360–402. doi: 10.1128/br.40.2.360-402.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boschwitz H., Milner Y., Keynan A., Halvorson H. O., Troll W. Effect of inhibitors of trypsin-like proteolytic enzymes Bacillus cereus T spore germination. J Bacteriol. 1983 Feb;153(2):700–708. doi: 10.1128/jb.153.2.700-708.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bulla L. A., Jr, Kramer K. J., Cox D. J., Jones B. L., Davidson L. I., Lookhart G. L. Purification and characterization of the entomocidal protoxin of Bacillus thuringiensis. J Biol Chem. 1981 Mar 25;256(6):3000–3004. [PubMed] [Google Scholar]
  5. GOLLAKOTA K. G., HALVORSON H. O. Biochemical changes occurring during sporulation of Bacillus cereus. Inhibition of sporulation by alpha-picolinic acid. J Bacteriol. 1960 Jan;79:1–8. doi: 10.1128/jb.79.1.1-8.1960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. HALMANN M., KEYNAN A. Stages in germination of spores of Bacillus licheniformis. J Bacteriol. 1962 Dec;84:1187–1193. doi: 10.1128/jb.84.6.1187-1193.1962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. HILLS G. M. Chemical factors in the germination of spore-bearing aerobes: observations on the influence of species, strain and conditions of growth. J Gen Microbiol. 1950 Jan;4(1):38–47. doi: 10.1099/00221287-4-1-38. [DOI] [PubMed] [Google Scholar]
  8. Jenkinson H. F. Altered arrangement of proteins in the spore coat of a germination mutant of Bacillus subtilis. J Gen Microbiol. 1983 Jun;129(6):1945–1958. doi: 10.1099/00221287-129-6-1945. [DOI] [PubMed] [Google Scholar]
  9. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  10. LUFT J. H. Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol. 1961 Feb;9:409–414. doi: 10.1083/jcb.9.2.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  12. Levinson H. S., Hyatt M. T. Sequence of events during Bacillus megaterim spore germination. J Bacteriol. 1966 May;91(5):1811–1818. doi: 10.1128/jb.91.5.1811-1818.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Loshon C. A., Swerdlow B. M., Setlow P. Bacillus megaterium spore protease. Synthesis and processing of precursor forms during sporulation and germination. J Biol Chem. 1982 Sep 25;257(18):10838–10845. [PubMed] [Google Scholar]
  14. Maurizi M. R., Switzer R. L. Proteolysis in bacterial sporulation. Curr Top Cell Regul. 1980;16:163–224. doi: 10.1016/b978-0-12-152816-4.50010-8. [DOI] [PubMed] [Google Scholar]
  15. Millet J. Characterization of a protein inhibitor of intracellular protease from Bacillus subtilis. FEBS Lett. 1977 Feb 15;74(1):59–61. doi: 10.1016/0014-5793(77)80752-7. [DOI] [PubMed] [Google Scholar]
  16. Moir A. Germination properties of a spore coat-defective mutant of Bacillus subtilis. J Bacteriol. 1981 Jun;146(3):1106–1116. doi: 10.1128/jb.146.3.1106-1116.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rossignol D. P., Vary J. C. A unique method for studying the initiation of Bacillus megaterium spore germination. Biochem Biophys Res Commun. 1977 Dec 21;79(4):1098–1103. doi: 10.1016/0006-291x(77)91118-4. [DOI] [PubMed] [Google Scholar]
  18. Srivastava O. P., Aronson A. I. Isolation and characterization of a unique protease from sporulating cells of Bacillus subtilis. Arch Microbiol. 1981 May;129(3):227–232. doi: 10.1007/BF00425256. [DOI] [PubMed] [Google Scholar]
  19. Tesone C., Torriani A. Protease associated with spores of Bacillus cereus. J Bacteriol. 1975 Oct;124(1):593–594. doi: 10.1128/jb.124.1.593-594.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Vary J. C., Kornberg A. Biochemical studies of bacterial sporulation and germination. XXI. Temperature-sensitive mutants for initiation of germination. J Bacteriol. 1970 Jan;101(1):327–329. doi: 10.1128/jb.101.1.327-329.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES