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. 1988 Oct;170(10):4727–4731. doi: 10.1128/jb.170.10.4727-4731.1988

Expression of the Saccharomyces cerevisiae PIS gene and synthesis of phosphatidylinositol in Escherichia coli.

J Nikawa 1, T Kodaki 1, S Yamashita 1
PMCID: PMC211514  PMID: 2844726

Abstract

Expression of the Saccharomyces cerevisiae PIS gene encoding phosphatidylinositol synthase in Escherichia coli was achieved by inserting its coding sequence into lacZ on pUC8. The fused gene encoded a phosphatidylinositol synthase whose amino-terminal three amino acids had been replaced by the amino-terminal five amino acids of E. coli beta-galactosidase. E. coli cells bearing this recombinant plasmid produced a significant level of phosphatidylinositol synthase in the presence of a lacZ inducer, isopropylthio-beta-D-galactopyranoside. When the culture medium was supplemented with myo-inositol and isopropylthio-beta-D-galactopyranoside, the cells accumulated a substantial amount of phosphatidylinositol in their membranes. When a saturating level of myo-inositol was added, phosphatidylinositol constituted about 4% of the total phospholipids. Phosphatidylinositol accumulation occurred at the expense of phosphatidylglycerol. The ratio of phosphatidylethanolamine to total acidic phospholipids remained constant. The growth rate of phosphatidylinositol-containing E. coli cells did not differ significantly from that of cells with the normal phospholipid composition.

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Selected References

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

  1. Ames G. F. Lipids of Salmonella typhimurium and Escherichia coli: structure and metabolism. J Bacteriol. 1968 Mar;95(3):833–843. doi: 10.1128/jb.95.3.833-843.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. BALLOU C. E., VILKAS E., LEDERER E. Structural studies on the myo-inositol phospholipids of Mycobacterium tuberculosis (var. bovis, strain BCG). J Biol Chem. 1963 Jan;238:69–76. [PubMed] [Google Scholar]
  3. Berridge M. J. Inositol trisphosphate and diacylglycerol as second messengers. Biochem J. 1984 Jun 1;220(2):345–360. doi: 10.1042/bj2200345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berridge M. J., Irvine R. F. Inositol trisphosphate, a novel second messenger in cellular signal transduction. Nature. 1984 Nov 22;312(5992):315–321. doi: 10.1038/312315a0. [DOI] [PubMed] [Google Scholar]
  5. Bolivar F., Backman K. Plasmids of Escherichia coli as cloning vectors. Methods Enzymol. 1979;68:245–267. doi: 10.1016/0076-6879(79)68018-7. [DOI] [PubMed] [Google Scholar]
  6. Cronan J. E., Vagelos P. R. Metabolism and function of the membrane phospholipids of Escherichia coli. Biochim Biophys Acta. 1972 Feb 14;265(1):25–60. doi: 10.1016/0304-4157(72)90018-4. [DOI] [PubMed] [Google Scholar]
  7. DITTMER J. C., LESTER R. L. A SIMPLE, SPECIFIC SPRAY FOR THE DETECTION OF PHOSPHOLIPIDS ON THIN-LAYER CHROMATOGRAMS. J Lipid Res. 1964 Jan;5:126–127. [PubMed] [Google Scholar]
  8. HUSTON C. K., ALBRO P. W., GRINDEY G. B. LIPIDS OF SARCINA LUTEA. 3. COMPOSITION OF THE COMPLEX LIPIDS. J Bacteriol. 1965 Mar;89:768–775. doi: 10.1128/jb.89.3.768-775.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hubbard S. C., Brody S. Glycerophospholipid variation in choline and inositol auxotrophs of Neurospora crassa. Internal compensation among zwitterionic and anionic species. J Biol Chem. 1975 Sep 25;250(18):7173–7181. [PubMed] [Google Scholar]
  10. Ikawa M. Bacterial phosphatides and natural relationships. Bacteriol Rev. 1967 Mar;31(1):54–64. doi: 10.1128/br.31.1.54-64.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kaibuchi K., Miyajima A., Arai K., Matsumoto K. Possible involvement of RAS-encoded proteins in glucose-induced inositolphospholipid turnover in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1986 Nov;83(21):8172–8176. doi: 10.1073/pnas.83.21.8172. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kates M. Bacterial lipids. Adv Lipid Res. 1964;2:17–90. [PubMed] [Google Scholar]
  13. Lester R. L., Steiner M. R. The occurrence of diphosphoinositide and triphosphoinositide in Saccharomyces cerevisiae. J Biol Chem. 1968 Sep 25;243(18):4889–4893. [PubMed] [Google Scholar]
  14. Michell R. H. Inositol phospholipids and cell surface receptor function. Biochim Biophys Acta. 1975 Mar 25;415(1):81–47. doi: 10.1016/0304-4157(75)90017-9. [DOI] [PubMed] [Google Scholar]
  15. Nikawa J., Kodaki T., Yamashita S. Primary structure and disruption of the phosphatidylinositol synthase gene of Saccharomyces cerevisiae. J Biol Chem. 1987 Apr 5;262(10):4876–4881. [PubMed] [Google Scholar]
  16. Nikawa J., Nagumo T., Yamashita S. Myo-inositol transport in Saccharomyces cerevisiae. J Bacteriol. 1982 May;150(2):441–446. doi: 10.1128/jb.150.2.441-446.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Nikawa J., Tsukagoshi Y., Kodaki T., Yamashita S. Nucleotide sequence and characterization of the yeast PSS gene encoding phosphatidylserine synthase. Eur J Biochem. 1987 Aug 17;167(1):7–12. doi: 10.1111/j.1432-1033.1987.tb13297.x. [DOI] [PubMed] [Google Scholar]
  18. Nikawa J., Yamashita S. Molecular cloning of the gene encoding CDPdiacylglycerol-inositol 3-phosphatidyl transferase in Saccharomyces cerevisiae. Eur J Biochem. 1984 Sep 3;143(2):251–256. doi: 10.1111/j.1432-1033.1984.tb08366.x. [DOI] [PubMed] [Google Scholar]
  19. Nikawa J., Yamashita S. Yeast mutant defective in synthesis of phosphatidylinositol. Isolation and characterization of a CDPdiacylglycerol--inositol 3-phosphatidyltransferase Km mutant. Eur J Biochem. 1982 Jul;125(2):445–451. doi: 10.1111/j.1432-1033.1982.tb06703.x. [DOI] [PubMed] [Google Scholar]
  20. Nishizuka Y. Turnover of inositol phospholipids and signal transduction. Science. 1984 Sep 21;225(4668):1365–1370. doi: 10.1126/science.6147898. [DOI] [PubMed] [Google Scholar]
  21. PATTERSON M. S., GREENE R. C. MEASUREMENT OF LOW ENERGY BETA-EMITTERS IN AQUEOUS SOLUTION BY LIQUID SCINTILLATION COUNTING OF EMULSIONS. Anal Chem. 1965 Jun;37:854–857. doi: 10.1021/ac60226a017. [DOI] [PubMed] [Google Scholar]
  22. Raetz C. R., Larson T. J., Dowhan W. Gene cloning for the isolation of enzymes of membrane lipid synthesis: phosphatidylserine synthase overproduction in Escherichia coli. Proc Natl Acad Sci U S A. 1977 Apr;74(4):1412–1416. doi: 10.1073/pnas.74.4.1412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Randle C. L., Albro P. W., Dittmer J. C. The phosphoglyceride composition of Gram-negative bacteria and the changes in composition during growth. Biochim Biophys Acta. 1969;187(2):214–220. doi: 10.1016/0005-2760(69)90030-7. [DOI] [PubMed] [Google Scholar]
  24. 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]
  25. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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