Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1988 Feb;170(2):775–780. doi: 10.1128/jb.170.2.775-780.1988

Disruption of the Escherichia coli cls gene responsible for cardiolipin synthesis.

S Nishijima 1, Y Asami 1, N Uetake 1, S Yamagoe 1, A Ohta 1, I Shibuya 1
PMCID: PMC210721  PMID: 2828323

Abstract

The cls gene of Escherichia coli is responsible for the synthesis of a major membrane phospholipid, cardiolipin, and has been proposed to encode cardiolipin synthase. This gene cloned on a pBR322 derivative was disrupted by either insertion of or replacement with a kanamycin-resistant gene followed by exchange with the homologous chromosomal region. The proper genomic disruptions were confirmed by Southern blot hybridization and a transductional linkage analysis. Both types of disruptants had essentially the same properties; cardiolipin synthase activity was not detectable, but the strains grew well, although their growth rates and final culture densities were lower than those of the corresponding wild-type strains and strains with the classical cls-1 mutation. A disruptant harboring a plasmid that carried the intact cls gene grew normally. The results indicate that the cls gene and probably the cardiolipin synthase are dispensable for E. coli but may confer growth or survival advantages. Low but definite levels of cardiolipin were synthesized by all the disruptants. Cardiolipin content of the cls mutants depended on the dosage of the pss gene, and attempts to transfer a null allele of the cls gene into a pss-1 mutant were unsuccessful. We point out the possibilities of minor cardiolipin formation by phosphatidylserine synthase and of the essential nature of cardiolipin for the survival of E. coli cells.

Full text

PDF
775

Images in this article

Selected References

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

  1. Bachmann B. J. Linkage map of Escherichia coli K-12, edition 7. Microbiol Rev. 1983 Jun;47(2):180–230. doi: 10.1128/mr.47.2.180-230.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  3. Casadaban M. J., Chou J., Cohen S. N. In vitro gene fusions that join an enzymatically active beta-galactosidase segment to amino-terminal fragments of exogenous proteins: Escherichia coli plasmid vectors for the detection and cloning of translational initiation signals. J Bacteriol. 1980 Aug;143(2):971–980. doi: 10.1128/jb.143.2.971-980.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Hirschberg C. B., Kennedy E. P. Mechanism of the enzymatic synthesis of cardiolipin in Escherichia coli. Proc Natl Acad Sci U S A. 1972 Mar;69(3):648–651. doi: 10.1073/pnas.69.3.648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Larson T. J., Dowhan W. Ribosomal-associated phosphatidylserine synthetase from Escherichia coli: purification by substrate-specific elution from phosphocellulose using cytidine 5'-diphospho-1,2-diacyl-sn-glycerol. Biochemistry. 1976 Nov 30;15(24):5212–5218. doi: 10.1021/bi00669a003. [DOI] [PubMed] [Google Scholar]
  6. Lovett M. A., Helinski D. R. Method for the isolation of the replication region of a bacterial replicon: construction of a mini-F'kn plasmid. J Bacteriol. 1976 Aug;127(2):982–987. doi: 10.1128/jb.127.2.982-987.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Miyazaki C., Kuroda M., Ohta A., Shibuya I. Genetic manipulation of membrane phospholipid composition in Escherichia coli: pgsA mutants defective in phosphatidylglycerol synthesis. Proc Natl Acad Sci U S A. 1985 Nov;82(22):7530–7534. doi: 10.1073/pnas.82.22.7530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ohta A., Obara T., Asami Y., Shibuya I. Molecular cloning of the cls gene responsible for cardiolipin synthesis in Escherichia coli and phenotypic consequences of its amplification. J Bacteriol. 1985 Aug;163(2):506–514. doi: 10.1128/jb.163.2.506-514.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ohta A., Shibuya I. Membrane phospholipid synthesis and phenotypic correlation of an Escherichia coli pss mutant. J Bacteriol. 1977 Nov;132(2):434–443. doi: 10.1128/jb.132.2.434-443.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Ohta A., Waggoner K., Louie K., Dowhan W. Cloning of genes involved in membrane lipid synthesis. Effects of amplification of phosphatidylserine synthase in Escherichia coli. J Biol Chem. 1981 Mar 10;256(5):2219–2225. [PubMed] [Google Scholar]
  11. Ono Y., White D. C. Cardiolipin-specific phospholipase D activity in Haemophilus parainfluenzae. J Bacteriol. 1970 Jul;103(1):111–115. doi: 10.1128/jb.103.1.111-115.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Pluschke G., Hirota Y., Overath P. Function of phospholipids in Escherichia coli. Characterization of a mutant deficient in cardiolipin synthesis. J Biol Chem. 1978 Jul 25;253(14):5048–5055. [PubMed] [Google Scholar]
  13. Pluschke G., Overath P. Function of phospholipids in Escherichia coli. Influence of changes in polar head group composition on the lipid phase transition and characterization of a mutant containing only saturated phospholipid acyl chains. J Biol Chem. 1981 Apr 10;256(7):3207–3212. [PubMed] [Google Scholar]
  14. Raetz C. R., Foulds J. Envelope composition and antibiotic hypersensitivity of Escherichia coli mutants defective in phosphatidylserine synthetase. J Biol Chem. 1977 Aug 25;252(16):5911–5915. [PubMed] [Google Scholar]
  15. Russel M., Model P. Replacement of the fip gene of Escherichia coli by an inactive gene cloned on a plasmid. J Bacteriol. 1984 Sep;159(3):1034–1039. doi: 10.1128/jb.159.3.1034-1039.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Shibuya I., Miyazaki C., Ohta A. Alteration of phospholipid composition by combined defects in phosphatidylserine and cardiolipin synthases and physiological consequences in Escherichia coli. J Bacteriol. 1985 Mar;161(3):1086–1092. doi: 10.1128/jb.161.3.1086-1092.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Shibuya I., Yamagoe S., Miyazaki C., Matsuzaki H., Ohta A. Biosynthesis of novel acidic phospholipid analogs in Escherichia coli. J Bacteriol. 1985 Feb;161(2):473–477. doi: 10.1128/jb.161.2.473-477.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Tunaitis E., Cronan J. E., Jr Characterization of the cardiolipin synthetase activity of Escherichia coli envelopes. Arch Biochem Biophys. 1973 Apr;155(2):420–427. doi: 10.1016/0003-9861(73)90132-x. [DOI] [PubMed] [Google Scholar]
  19. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  20. Yamaguchi K., Tomizawa J. Establishment of Escherichia coli cells with an integrated high copy number plasmid. Mol Gen Genet. 1980;178(3):525–533. doi: 10.1007/BF00337857. [DOI] [PubMed] [Google Scholar]
  21. van den Bosch H. Phosphoglyceride metabolism. Annu Rev Biochem. 1974;43(0):243–277. doi: 10.1146/annurev.bi.43.070174.001331. [DOI] [PubMed] [Google Scholar]

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

RESOURCES