• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jbacterPermissionsJournals.ASM.orgJournalJB ArticleJournal InfoAuthorsReviewers
J Bacteriol. Jan 1995; 177(2): 326–335.
PMCID: PMC176595

Cloning, nucleotide sequence, and mutagenesis of the Bacillus subtilis ponA operon, which codes for penicillin-binding protein (PBP) 1 and a PBP-related factor.

Abstract

An oligonucleotide probe designed to hybridize to genes encoding class A high-molecular-weight penicillin-binding proteins (PBPs) was used to identify the ponA gene encoding PBP1a and -1b (PBP1) of Bacillus subtilis. The identity of the ponA product was established by (i) the presence of a sequence coding for a peptide generated from PBP1 and (ii) the disappearance of PBP1 in a ponA mutant. DNA sequence analysis revealed that the amino acid sequence of PBP1 was similar to those of other class A high-molecular-weight PBPs and that ponA appeared to be cotranscribed with an upstream gene (termed prfA) of unknown function. Null mutations in ponA resulted in a slight decrease in growth rate and a change in colony morphology but had no significant effect on cell morphology, cell division, sporulation, spore heat resistance, or spore germination. Mutations in prfA which did not effect ponA expression produced a more significant decrease in growth rate but had no other significant phenotypic effects. Deletion of both prfA and ponA resulted in extremely slow growth and a reduction in sporulation efficiency. Studies of expression of transcriptional fusions of ponA and prfA to lacZ demonstrated that these two genes constitute an operon. Expression of these genes was relatively constant during growth, decreased during sporulation, and was induced approximately 15 min into spore germination. The ponA locus was mapped to the 200 degrees region of the chromosomal physical map.

Full Text

The Full Text of this article is available as a PDF (509K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Adachi H, Ohta T, Matsuzawa H. A water-soluble form of penicillin-binding protein 2 of Escherichia coli constructed by site-directed mutagenesis. FEBS Lett. 1987 Dec 21;226(1):150–154. [PubMed]
  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. [PubMed]
  • 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. [PMC free article] [PubMed]
  • Blumberg PM, Strominger JL. Five penicillin-binding components occur in Bacillus subtilis membranes. J Biol Chem. 1972 Dec 25;247(24):8107–8113. [PubMed]
  • Broome-Smith JK, Edelman A, Yousif S, Spratt BG. The nucleotide sequences of the ponA and ponB genes encoding penicillin-binding protein 1A and 1B of Escherichia coli K12. Eur J Biochem. 1985 Mar 1;147(2):437–446. [PubMed]
  • Buchanan CE, 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. [PMC free article] [PubMed]
  • Buchanan CE, Ling ML. 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. [PMC free article] [PubMed]
  • Buchanan CE, Strominger JL. Altered penicillin-binding components in penicillin-resistant mutants of Bacillus subtilis. Proc Natl Acad Sci U S A. 1976 Jun;73(6):1816–1820. [PMC free article] [PubMed]
  • Daniel RA, Drake S, Buchanan CE, 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. [PubMed]
  • Edelman A, Bowler L, Broome-Smith JK, Spratt BG. Use of a beta-lactamase fusion vector to investigate the organization of penicillin-binding protein 1B in the cytoplasmic membrane of Escherichia coli. Mol Microbiol. 1987 Jul;1(1):101–106. [PubMed]
  • Ferrari FA, Nguyen A, Lang D, Hoch JA. Construction and properties of an integrable plasmid for Bacillus subtilis. J Bacteriol. 1983 Jun;154(3):1513–1515. [PMC free article] [PubMed]
  • Ghuysen JM. Serine beta-lactamases and penicillin-binding proteins. Annu Rev Microbiol. 1991;45:37–67. [PubMed]
  • Hansson M, Hederstedt L. Cloning and characterization of the Bacillus subtilis hemEHY gene cluster, which encodes protoheme IX biosynthetic enzymes. J Bacteriol. 1992 Dec;174(24):8081–8093. [PMC free article] [PubMed]
  • Hara H, Nishimura Y, Kato J, Suzuki H, Nagasawa H, Suzuki A, Hirota Y. Genetic analyses of processing involving C-terminal cleavage in penicillin-binding protein 3 of Escherichia coli. J Bacteriol. 1989 Nov;171(11):5882–5889. [PMC free article] [PubMed]
  • Ishino F, Mitsui K, Tamaki S, Matsuhashi M. Dual enzyme activities of cell wall peptidoglycan synthesis, peptidoglycan transglycosylase and penicillin-sensitive transpeptidase, in purified preparations of Escherichia coli penicillin-binding protein 1A. Biochem Biophys Res Commun. 1980 Nov 17;97(1):287–293. [PubMed]
  • Itaya M, Tanaka T. Complete physical map of the Bacillus subtilis 168 chromosome constructed by a gene-directed mutagenesis method. J Mol Biol. 1991 Aug 5;220(3):631–648. [PubMed]
  • Jacques P, el Kharroubi A, Van Beeumen J, Piras G, Coyette J, Ghuysen JM. Mode of membrane insertion and sequence of a 32-amino acid peptide stretch of the penicillin-binding protein 4 of Enterococcus hirae. FEMS Microbiol Lett. 1991 Aug 1;66(2):119–123. [PubMed]
  • Kleppe G, Yu W, Strominger JL. Penicillin-binding proteins in Bacillus subtilis mutants. Antimicrob Agents Chemother. 1982 Jun;21(6):979–983. [PMC free article] [PubMed]
  • LeDeaux JR, Grossman AD. 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. [PMC free article] [PubMed]
  • Leighton TJ, Doi RH. The stability of messenger ribonucleic acid during sporulation in Bacillus subtilis. J Biol Chem. 1971 May 25;246(10):3189–3195. [PubMed]
  • LOWRY OH, ROSEBROUGH NJ, FARR AL, RANDALL RJ. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed]
  • Martin C, Briese T, Hakenbeck R. Nucleotide sequences of genes encoding penicillin-binding proteins from Streptococcus pneumoniae and Streptococcus oralis with high homology to Escherichia coli penicillin-binding proteins 1a and 1b. J Bacteriol. 1992 Jul;174(13):4517–4523. [PMC free article] [PubMed]
  • Nagasawa H, Sakagami Y, Suzuki A, Suzuki H, Hara H, Hirota Y. Determination of the cleavage site involved in C-terminal processing of penicillin-binding protein 3 of Escherichia coli. J Bacteriol. 1989 Nov;171(11):5890–5893. [PMC free article] [PubMed]
  • Neyman SL, Buchanan CE. 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. [PMC free article] [PubMed]
  • Nicholas RA, Lamson DR, Schultz DE. Penicillin-binding protein 1B from Escherichia coli contains a membrane association site in addition to its transmembrane anchor. J Biol Chem. 1993 Mar 15;268(8):5632–5641. [PubMed]
  • Popham DL, 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. [PMC free article] [PubMed]
  • Popham DL, 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. [PMC free article] [PubMed]
  • Popham DL, 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. [PMC free article] [PubMed]
  • Sowell MO, Buchanan CE. Changes in penicillin-binding proteins during sporulation of Bacillus subtilis. J Bacteriol. 1983 Mar;153(3):1331–1337. [PMC free article] [PubMed]
  • Suzuki H, van Heijenoort Y, Tamura T, Mizoguchi J, Hirota Y, van Heijenoort J. In vitro peptidoglycan polymerization catalysed by penicillin binding protein 1b of Escherichia coli K-12. FEBS Lett. 1980 Feb 11;110(2):245–249. [PubMed]
  • Suzuki H, Nishimura Y, Hirota Y. On the process of cellular division in Escherichia coli: a series of mutants of E. coli altered in the penicillin-binding proteins. Proc Natl Acad Sci U S A. 1978 Feb;75(2):664–668. [PMC free article] [PubMed]
  • Todd JA, Roberts AN, Johnstone K, Piggot PJ, Winter G, Ellar DJ. 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. [PMC free article] [PubMed]
  • von Heijne G. Transcending the impenetrable: how proteins come to terms with membranes. Biochim Biophys Acta. 1988 Jun 9;947(2):307–333. [PubMed]
  • Weigel LM, Belisle JT, Radolf JD, Norgard MV. Digoxigenin-ampicillin conjugate for detection of penicillin-binding proteins by chemiluminescence. Antimicrob Agents Chemother. 1994 Feb;38(2):330–336. [PMC free article] [PubMed]
  • Wu JJ, Schuch R, Piggot PJ. 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. [PMC free article] [PubMed]
  • Yanouri A, Daniel RA, Errington J, Buchanan CE. 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. [PMC free article] [PubMed]
  • Yoshikawa H. DNA synthesis during germination of Bacillus subtilis spores. Proc Natl Acad Sci U S A. 1965 Jun;53(6):1476–1483. [PMC free article] [PubMed]
  • Yousif SY, Broome-Smith JK, Spratt BG. Lysis of Escherichia coli by beta-lactam antibiotics: deletion analysis of the role of penicillin-binding proteins 1A and 1B. J Gen Microbiol. 1985 Oct;131(10):2839–2845. [PubMed]

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

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...