Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Oct;178(20):6001–6005. doi: 10.1128/jb.178.20.6001-6005.1996

Identification and characterization of pbpC, the gene encoding Bacillus subtilis penicillin-binding protein 3.

T Murray 1, D L Popham 1, P Setlow 1
PMCID: PMC178458  PMID: 8830698

Abstract

Penicillin-binding proteins (PBPs) are enzymes involved in the synthesis of peptidoglycan structures in Bacillus subtilis such as the vegetative cell wall and the spore cortex. The B. subtilis sequencing project has identified a gene (orf16, EMBL accession number D38161) which exhibits significant sequence similarity to genes encoding class B high-molecular-weight PBPs. We have found that orf16 encodes PBP3 and have renamed this locus pbpC. Transcriptional fusions to lacZ were used to demonstrate that pbpC is transcribed primarily during log-phase growth, with lower amounts expressed during sporulation. During spore germination and outgrowth, pbpC expression resumes coincident with an increase in the optical density of the culture. The major promoter for pbpC is located just upstream of the gene; a low level of expression during sporulation appears to originate from much further upstream. Loss of PBP3 does not produce any detectable change in phenotype with respect to cell morphology, growth, sporulation, spore heat resistance, or spore germination and outgrowth. This was also true when the pbpC mutation was combined with mutations affecting other PBP-encoding genes to produce double mutants. These findings are consistent with previous evidence that many PBPs of B. subtilis have redundant functions within the cell.

Full Text

The Full Text of this article is available as a PDF (365.2 KB).

Selected References

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

  1. Adachi H., Ishiguro M., Imajoh S., Ohta T., Matsuzawa H. Active-site residues of the transpeptidase domain of penicillin-binding protein 2 from Escherichia coli: similarity in catalytic mechanism to class A beta-lactamases. Biochemistry. 1992 Jan 21;31(2):430–437. doi: 10.1021/bi00117a018. [DOI] [PubMed] [Google Scholar]
  2. Akagawa E., Kurita K., Sugawara T., Nakamura K., Kasahara Y., Ogasawara N., Yamane K. Determination of a 17,484 bp nucleotide sequence around the 39 degrees region of the Bacillus subtilis chromosome and similarity analysis of the products of putative ORFs. Microbiology. 1995 Dec;141(Pt 12):3241–3245. doi: 10.1099/13500872-141-12-3241. [DOI] [PubMed] [Google Scholar]
  3. Antoniewski C., Savelli B., Stragier P. The spoIIJ gene, which regulates early developmental steps in Bacillus subtilis, belongs to a class of environmentally responsive genes. J Bacteriol. 1990 Jan;172(1):86–93. doi: 10.1128/jb.172.1.86-93.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blumberg P. M., Strominger J. L. Five penicillin-binding components occur in Bacillus subtilis membranes. J Biol Chem. 1972 Dec 25;247(24):8107–8113. [PubMed] [Google Scholar]
  5. Buchanan C. E., Gustafson A. Mutagenesis and mapping of the gene for a sporulation-specific penicillin-binding protein in Bacillus subtilis. J Bacteriol. 1992 Aug;174(16):5430–5435. doi: 10.1128/jb.174.16.5430-5435.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Buchanan C. E., Ling M. L. Isolation and sequence analysis of dacB, which encodes a sporulation-specific penicillin-binding protein in Bacillus subtilis. J Bacteriol. 1992 Mar;174(6):1717–1725. doi: 10.1128/jb.174.6.1717-1725.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Daniel R. A., Drake S., Buchanan C. E., Scholle R., Errington J. The Bacillus subtilis spoVD gene encodes a mother-cell-specific penicillin-binding protein required for spore morphogenesis. J Mol Biol. 1994 Jan 7;235(1):209–220. doi: 10.1016/s0022-2836(05)80027-0. [DOI] [PubMed] [Google Scholar]
  8. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ghuysen J. M. Serine beta-lactamases and penicillin-binding proteins. Annu Rev Microbiol. 1991;45:37–67. doi: 10.1146/annurev.mi.45.100191.000345. [DOI] [PubMed] [Google Scholar]
  10. Hartman B. J., Tomasz A. Low-affinity penicillin-binding protein associated with beta-lactam resistance in Staphylococcus aureus. J Bacteriol. 1984 May;158(2):513–516. doi: 10.1128/jb.158.2.513-516.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kleppe G., Strominger J. L. Studies of the high molecular weight penicillin-binding proteins of Bacillus subtilis. J Biol Chem. 1979 Jun 10;254(11):4856–4862. [PubMed] [Google Scholar]
  12. Kleppe G., Yu W., Strominger J. L. Penicillin-binding proteins in Bacillus subtilis mutants. Antimicrob Agents Chemother. 1982 Jun;21(6):979–983. doi: 10.1128/aac.21.6.979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. LeDeaux J. R., Grossman A. D. Isolation and characterization of kinC, a gene that encodes a sensor kinase homologous to the sporulation sensor kinases KinA and KinB in Bacillus subtilis. J Bacteriol. 1995 Jan;177(1):166–175. doi: 10.1128/jb.177.1.166-175.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Leighton T. J., Doi R. H. The stability of messenger ribonucleic acid during sporulation in Bacillus subtilis. J Biol Chem. 1971 May 25;246(10):3189–3195. [PubMed] [Google Scholar]
  15. Magni C., Marini P., de Mendoza D. Extraction of RNA from gram-positive bacteria. Biotechniques. 1995 Dec;19(6):880–884. [PubMed] [Google Scholar]
  16. Moszer I., Glaser P., Danchin A. SubtiList: a relational database for the Bacillus subtilis genome. Microbiology. 1995 Feb;141(Pt 2):261–268. doi: 10.1099/13500872-141-2-261. [DOI] [PubMed] [Google Scholar]
  17. Neyman S. L., Buchanan C. E. Restoration of vegetative penicillin-binding proteins during germination and outgrowth of Bacillus subtilis spores: relationship of individual proteins to specific cell cycle events. J Bacteriol. 1985 Jan;161(1):164–168. doi: 10.1128/jb.161.1.164-168.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Patel-King R. S., Benashki S. E., Harrison A., King S. M. Two functional thioredoxins containg redox-senesitive vicinal dithiols from the Chlamydomonas outer dynein arm. J Biol Chem. 1996 Mar 15;271(11):6283–6291. doi: 10.1074/jbc.271.11.6283. [DOI] [PubMed] [Google Scholar]
  19. Piras G., Raze D., el Kharroubi A., Hastir D., Englebert S., Coyette J., Ghuysen J. M. Cloning and sequencing of the low-affinity penicillin-binding protein 3r-encoding gene of Enterococcus hirae S185: modular design and structural organization of the protein. J Bacteriol. 1993 May;175(10):2844–2852. doi: 10.1128/jb.175.10.2844-2852.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Popham D. L., Illades-Aguiar B., Setlow P. The Bacillus subtilis dacB gene, encoding penicillin-binding protein 5*, is part of a three-gene operon required for proper spore cortex synthesis and spore core dehydration. J Bacteriol. 1995 Aug;177(16):4721–4729. doi: 10.1128/jb.177.16.4721-4729.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Popham D. L., Setlow P. Cloning, nucleotide sequence, and mutagenesis of the Bacillus subtilis ponA operon, which codes for penicillin-binding protein (PBP) 1 and a PBP-related factor. J Bacteriol. 1995 Jan;177(2):326–335. doi: 10.1128/jb.177.2.326-335.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Popham D. L., Setlow P. Cloning, nucleotide sequence, and regulation of the Bacillus subtilis pbpE operon, which codes for penicillin-binding protein 4* and an apparent amino acid racemase. J Bacteriol. 1993 May;175(10):2917–2925. doi: 10.1128/jb.175.10.2917-2925.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Popham D. L., Setlow P. Cloning, nucleotide sequence, and regulation of the Bacillus subtilis pbpF gene, which codes for a putative class A high-molecular-weight penicillin-binding protein. J Bacteriol. 1993 Aug;175(15):4870–4876. doi: 10.1128/jb.175.15.4870-4876.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Popham D. L., Setlow P. Cloning, nucleotide sequence, mutagenesis, and mapping of the Bacillus subtilis pbpD gene, which codes for penicillin-binding protein 4. J Bacteriol. 1994 Dec;176(23):7197–7205. doi: 10.1128/jb.176.23.7197-7205.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Popham D. L., Setlow P. Phenotypes of Bacillus subtilis mutants lacking multiple class A high-molecular-weight penicillin-binding proteins. J Bacteriol. 1996 Apr;178(7):2079–2085. doi: 10.1128/jb.178.7.2079-2085.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Popham D. L., Setlow P. The cortical peptidoglycan from spores of Bacillus megaterium and Bacillus subtilis is not highly cross-linked. J Bacteriol. 1993 May;175(9):2767–2769. doi: 10.1128/jb.175.9.2767-2769.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Smith H., Bron S., Van Ee J., Venema G. Construction and use of signal sequence selection vectors in Escherichia coli and Bacillus subtilis. J Bacteriol. 1987 Jul;169(7):3321–3328. doi: 10.1128/jb.169.7.3321-3328.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Smith H., de Jong A., Bron S., Venema G. Characterization of signal-sequence-coding regions selected from the Bacillus subtilis chromosome. Gene. 1988 Oct 30;70(2):351–361. doi: 10.1016/0378-1119(88)90207-7. [DOI] [PubMed] [Google Scholar]
  30. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  31. Sowell M. O., Buchanan C. E. Changes in penicillin-binding proteins during sporulation of Bacillus subtilis. J Bacteriol. 1983 Mar;153(3):1331–1337. doi: 10.1128/jb.153.3.1331-1337.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Sun D. X., Cabrera-Martinez R. M., Setlow P. Control of transcription of the Bacillus subtilis spoIIIG gene, which codes for the forespore-specific transcription factor sigma G. J Bacteriol. 1991 May;173(9):2977–2984. doi: 10.1128/jb.173.9.2977-2984.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Todd J. A., Roberts A. N., Johnstone K., Piggot P. J., Winter G., Ellar D. J. Reduced heat resistance of mutant spores after cloning and mutagenesis of the Bacillus subtilis gene encoding penicillin-binding protein 5. J Bacteriol. 1986 Jul;167(1):257–264. doi: 10.1128/jb.167.1.257-264.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wu J. J., Schuch R., Piggot P. J. Characterization of a Bacillus subtilis sporulation operon that includes genes for an RNA polymerase sigma factor and for a putative DD-carboxypeptidase. J Bacteriol. 1992 Aug;174(15):4885–4892. doi: 10.1128/jb.174.15.4885-4892.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Yanouri A., Daniel R. A., Errington J., Buchanan C. E. Cloning and sequencing of the cell division gene pbpB, which encodes penicillin-binding protein 2B in Bacillus subtilis. J Bacteriol. 1993 Dec;175(23):7604–7616. doi: 10.1128/jb.175.23.7604-7616.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES