Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1993 May;175(9):2568–2577. doi: 10.1128/jb.175.9.2568-2577.1993

Proteolytic processing of the protease which initiates degradation of small, acid-soluble proteins during germination of Bacillus subtilis spores.

J L Sanchez-Salas 1, P Setlow 1
PMCID: PMC204558  PMID: 8478323

Abstract

Degradation of small, acid-soluble spore proteins during germination of Bacillus subtilis spores is initiated by a sequence-specific protease called GPR. Western blot (immunoblot) analysis of either Bacillus megaterium or B. subtilis GPR expressed in B. subtilis showed that GPR is synthesized at about the third hour of sporulation in a precursor form and is processed to an approximately 2- to 5-kDa-smaller species 2 to 3 h later, at or slightly before the time of accumulation of dipicolinic acid by the forespore. This was found with both normal levels of expression of B. subtilis and B. megaterium GPR in B. subtilis, as well as when either protein was overexpressed up to 100-fold. The sporulation-specific processing of GPR was blocked in all spoIII, -IV, and -V mutants tested (none of which accumulated dipicolinic acid), but not in a spoVI mutant which accumulated dipicolinic acid. The amino-terminal sequences of the B. megaterium and B. subtilis GPR initially synthesized in sporulation were identical to those predicted from the coding genes' sequences. However, the processed form generated in sporulation lacked 15 (B. megaterium) or 16 (B. subtilis) amino-terminal residues. The amino acid sequence surrounding this proteolytic cleavage site was very homologous to the consensus sequence recognized and cleaved by GPR in its small, acid-soluble spore protein substrates. This observation, plus the efficient processing of overproduced GPR during sporulation, suggests that the GPR precursor may autoproteolyze itself during sporulation. During spore germination, the GPR from either species expressed in B. subtilis was further processed by removal of one additional amino-terminal amino acid (leucine), generating the mature protease which acts during spore germination.

Full text

PDF
2568

Images in this article

2571Image
on p.2571
2571Image
on p.2571
2572Image
on p.2572
2572Image
on p.2572
2574Image
on p.2574

Selected References

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

  1. Bachmair A., Finley D., Varshavsky A. In vivo half-life of a protein is a function of its amino-terminal residue. Science. 1986 Oct 10;234(4773):179–186. doi: 10.1126/science.3018930. [DOI] [PubMed] [Google Scholar]
  2. Connors M. J., Setlow P. Cloning of a small, acid-soluble spore protein gene from Bacillus subtilis and determination of its complete nucleotide sequence. J Bacteriol. 1985 Jan;161(1):333–339. doi: 10.1128/jb.161.1.333-339.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cutting S., Oke V., Driks A., Losick R., Lu S., Kroos L. A forespore checkpoint for mother cell gene expression during development in B. subtilis. Cell. 1990 Jul 27;62(2):239–250. doi: 10.1016/0092-8674(90)90362-i. [DOI] [PubMed] [Google Scholar]
  4. Errington J., Cutting S. M., Mandelstam J. Branched pattern of regulatory interactions between late sporulation genes in Bacillus subtilis. J Bacteriol. 1988 Feb;170(2):796–801. doi: 10.1128/jb.170.2.796-801.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Goldrick S., Setlow P. Expression of a Bacillus megaterium sporulation-specific gene during sporulation of Bacillus subtilis. J Bacteriol. 1983 Sep;155(3):1459–1462. doi: 10.1128/jb.155.3.1459-1462.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hackett R. H., Setlow P. Enzymatic activity of precursors of Bacillus megaterium spore protease. J Bacteriol. 1983 Jan;153(1):375–378. doi: 10.1128/jb.153.1.375-378.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. LaBell T. L., Trempy J. E., Haldenwang W. G. Sporulation-specific sigma factor sigma 29 of Bacillus subtilis is synthesized from a precursor protein, P31. Proc Natl Acad Sci U S A. 1987 Apr;84(7):1784–1788. doi: 10.1073/pnas.84.7.1784. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Loshon C. A., Setlow P. Bacillus megaterium spore protease: purification, radioimmunoassay, and analysis of antigen level and localization during growth, sporulation, and spore germination. J Bacteriol. 1982 Apr;150(1):303–311. doi: 10.1128/jb.150.1.303-311.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. 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]
  10. Losick R., Stragier P. Crisscross regulation of cell-type-specific gene expression during development in B. subtilis. Nature. 1992 Feb 13;355(6361):601–604. doi: 10.1038/355601a0. [DOI] [PubMed] [Google Scholar]
  11. Lu S., Halberg R., Kroos L. Processing of the mother-cell sigma factor, sigma K, may depend on events occurring in the forespore during Bacillus subtilis development. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9722–9726. doi: 10.1073/pnas.87.24.9722. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mason J. M., Hackett R. H., Setlow P. Regulation of expression of genes coding for small, acid-soluble proteins of Bacillus subtilis spores: studies using lacZ gene fusions. J Bacteriol. 1988 Jan;170(1):239–244. doi: 10.1128/jb.170.1.239-244.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mason J. M., Setlow P. Different small, acid-soluble proteins of the alpha/beta type have interchangeable roles in the heat and UV radiation resistance of Bacillus subtilis spores. J Bacteriol. 1987 Aug;169(8):3633–3637. doi: 10.1128/jb.169.8.3633-3637.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Mason J. M., Setlow P. Essential role of small, acid-soluble spore proteins in resistance of Bacillus subtilis spores to UV light. J Bacteriol. 1986 Jul;167(1):174–178. doi: 10.1128/jb.167.1.174-178.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Nicholson W. L., Setlow P. Dramatic increase in negative superhelicity of plasmid DNA in the forespore compartment of sporulating cells of Bacillus subtilis. J Bacteriol. 1990 Jan;172(1):7–14. doi: 10.1128/jb.172.1.7-14.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sanchez-Salas J. L., Santiago-Lara M. L., Setlow B., Sussman M. D., Setlow P. Properties of Bacillus megaterium and Bacillus subtilis mutants which lack the protease that degrades small, acid-soluble proteins during spore germination. J Bacteriol. 1992 Feb;174(3):807–814. doi: 10.1128/jb.174.3.807-814.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Setlow P. Protease and peptidase activities in growing and sporulating cells and dormant spores of Bacillus megaterium. J Bacteriol. 1975 May;122(2):642–649. doi: 10.1128/jb.122.2.642-649.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Setlow P. Small, acid-soluble spore proteins of Bacillus species: structure, synthesis, genetics, function, and degradation. Annu Rev Microbiol. 1988;42:319–338. doi: 10.1146/annurev.mi.42.100188.001535. [DOI] [PubMed] [Google Scholar]
  19. Shimotsu H., Henner D. J. Construction of a single-copy integration vector and its use in analysis of regulation of the trp operon of Bacillus subtilis. Gene. 1986;43(1-2):85–94. doi: 10.1016/0378-1119(86)90011-9. [DOI] [PubMed] [Google Scholar]
  20. Stragier P., Bonamy C., Karmazyn-Campelli C. Processing of a sporulation sigma factor in Bacillus subtilis: how morphological structure could control gene expression. Cell. 1988 Mar 11;52(5):697–704. doi: 10.1016/0092-8674(88)90407-2. [DOI] [PubMed] [Google Scholar]
  21. Sun D. X., Setlow P. Cloning, nucleotide sequence, and expression of the Bacillus subtilis ans operon, which codes for L-asparaginase and L-aspartase. J Bacteriol. 1991 Jun;173(12):3831–3845. doi: 10.1128/jb.173.12.3831-3845.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sun D., Fajardo-Cavazos P., Sussman M. D., Tovar-Rojo F., Cabrera-Martinez R. M., Setlow P. Effect of chromosome location of Bacillus subtilis forespore genes on their spo gene dependence and transcription by E sigma F: identification of features of good E sigma F-dependent promoters. J Bacteriol. 1991 Dec;173(24):7867–7874. doi: 10.1128/jb.173.24.7867-7874.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Sussman M. D., Setlow P. Cloning, nucleotide sequence, and regulation of the Bacillus subtilis gpr gene, which codes for the protease that initiates degradation of small, acid-soluble proteins during spore germination. J Bacteriol. 1991 Jan;173(1):291–300. doi: 10.1128/jb.173.1.291-300.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Tobias J. W., Shrader T. E., Rocap G., Varshavsky A. The N-end rule in bacteria. Science. 1991 Nov 29;254(5036):1374–1377. doi: 10.1126/science.1962196. [DOI] [PubMed] [Google Scholar]
  25. Tovar-Rojo F., Setlow P. Effects of mutant small, acid-soluble spore proteins from Bacillus subtilis on DNA in vivo and in vitro. J Bacteriol. 1991 Aug;173(15):4827–4835. doi: 10.1128/jb.173.15.4827-4835.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES