Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1987 Jan;169(1):380–385. doi: 10.1128/jb.169.1.380-385.1987

Characterization of the specific pyruvate transport system in Escherichia coli K-12.

V J Lang, C Leystra-Lantz, R A Cook
PMCID: PMC211778  PMID: 3025181

Abstract

A mutant of Escherichia coli K-12 lacking pyruvate dehydrogenase and phosphoenolpyruvate synthase was used to study the transport of pyruvate by whole cells. Uptake of pyruvate was maximal in mid-log phase cells, with a Michaelis constant for transport of 20 microM. Pretreatment of the cells with respiratory chain poisons or uncouplers, except for arsenate, inhibited transport up to 95%. Lactate and alanine were competitive inhibitors, but at nonphysiological concentrations. The synthetic analogs 3-bromopyruvate and pyruvic acid methyl ester inhibited competitively. The uptake of pyruvate was also characterized in membrane vesicles from wild-type E. coli K-12. Transport required an artificial electron donor system, phenazine methosulfate and sodium ascorbate. Pyruvate was concentrated in vesicles 7- to 10-fold over the external concentration, with a Michaelis constant of 15 microM. Energy poisons, except arsenate, inhibited the transport of pyruvate. Synthetic analogs such as 3-bromopyruvate were competitive inhibitors of transport. Lactate initially appeared to be a competitive inhibitor of pyruvate transport in vesicles, but this was a result of oxidation of lactate to pyruvate. The results indicate that uptake of pyruvate in E. coli is via a specific active transport system.

Full text

PDF
380

Selected References

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

  1. Abendano J. J., Kepes A. Sensitization of D-glucuronic acid transport system of E. coli to protein group reagents in presence of substrate or absence of energy source. Biochem Biophys Res Commun. 1973 Oct 15;54(4):1342–1346. doi: 10.1016/0006-291x(73)91134-0. [DOI] [PubMed] [Google Scholar]
  2. Bewick M. A., Lo T. C. Localization of the dicarboxylate binding protein in the cell envelope of Escherichia coli K12. Can J Biochem. 1980 Oct;58(10):885–897. doi: 10.1139/o80-123. [DOI] [PubMed] [Google Scholar]
  3. Brockman R. W., Heppel L. A. On the localization of alkaline phosphatase and cyclic phosphodiesterase in Escherichia coli. Biochemistry. 1968 Jul;7(7):2554–2562. doi: 10.1021/bi00847a016. [DOI] [PubMed] [Google Scholar]
  4. Gibson J. Uptake of C4 dicarboxylates and pyruvate by Rhodopseudomonas spheroides. J Bacteriol. 1975 Aug;123(2):471–480. doi: 10.1128/jb.123.2.471-480.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Johnson C. L., Cha Y. A., Stern J. R. Citrate uptake in membrane vesicles of Klebsiella aerogenes. J Bacteriol. 1975 Feb;121(2):682–687. doi: 10.1128/jb.121.2.682-687.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Kaback H. R., Milner L. S. Relationship of a membrane-bound D-(-)-lactic dehydrogenase to amino acid transport in isolated bacterial membrane preparations. Proc Natl Acad Sci U S A. 1970 Jul;66(3):1008–1015. doi: 10.1073/pnas.66.3.1008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Kaback H. R., Stadtman E. R. Proline uptake by an isolated cytoplasmic membrane preparation of Escherichia coli. Proc Natl Acad Sci U S A. 1966 Apr;55(4):920–927. doi: 10.1073/pnas.55.4.920. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kay W. W., Cameron M. Citrate transport in Salmonella typhimurium. Arch Biochem Biophys. 1978 Sep;190(1):270–280. doi: 10.1016/0003-9861(78)90276-x. [DOI] [PubMed] [Google Scholar]
  9. Kay W. W. Genetic control of the metabolism of propionate by Escherichia coli K12. Biochim Biophys Acta. 1972 May 16;264(3):508–521. doi: 10.1016/0304-4165(72)90014-1. [DOI] [PubMed] [Google Scholar]
  10. Kay W. W., Kornberg H. L. The uptake of C4-dicarboxylic acids by Escherichia coli. Eur J Biochem. 1971 Jan;18(2):274–281. doi: 10.1111/j.1432-1033.1971.tb01240.x. [DOI] [PubMed] [Google Scholar]
  11. Kay W. W. Two aspartate transport systems in Escherichia coli. J Biol Chem. 1971 Dec 10;246(23):7373–7382. [PubMed] [Google Scholar]
  12. Konings W. N. Active transport of solutes in bacterial membrane vesicles. Adv Microb Physiol. 1977;15:175–251. doi: 10.1016/s0065-2911(08)60317-3. [DOI] [PubMed] [Google Scholar]
  13. Kornberg H. L., Smith J. Genetic control of the uptake of pyruvate by Escherichia coli. Biochim Biophys Acta. 1967 Nov 28;148(2):591–592. doi: 10.1016/0304-4165(67)90167-5. [DOI] [PubMed] [Google Scholar]
  14. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  15. Langley D., Guest J. R. Biochemical genetics of the alpha-keto acid dehydrogenase complexes of Escherichia coli K12: isolation and biochemical properties of deletion mutants. J Gen Microbiol. 1977 Apr;99(2):263–276. doi: 10.1099/00221287-99-2-263. [DOI] [PubMed] [Google Scholar]
  16. Lo T. C., Bewick M. A. Use of a nonpenetrating substrate analogue to study the molecular mechanism of the outer membrane dicarboxylate transport system in Escherichia coli K12. J Biol Chem. 1981 Jun 10;256(11):5511–5517. [PubMed] [Google Scholar]
  17. Lo T. C. The molecular mechanism of dicarboxylic acid transport in Escherichia coli K 12. J Supramol Struct. 1977;7(3-4):463–480. doi: 10.1002/jss.400070316. [DOI] [PubMed] [Google Scholar]
  18. Matin A., Konings W. N. Transport of lactate and succinate by membrane vesicles of Escherichia coli, Bacillus subtilis and a pseudomonas species. Eur J Biochem. 1973 Apr 2;34(1):58–67. doi: 10.1111/j.1432-1033.1973.tb02728.x. [DOI] [PubMed] [Google Scholar]
  19. Nossal N. G., Heppel L. A. The release of enzymes by osmotic shock from Escherichia coli in exponential phase. J Biol Chem. 1966 Jul 10;241(13):3055–3062. [PubMed] [Google Scholar]
  20. Nygaard P. Two-way separation of carboxylic acids by thin layer electrophoresis and chromatography. J Chromatogr. 1967 Sep;30(1):240–243. doi: 10.1016/s0021-9673(00)84145-x. [DOI] [PubMed] [Google Scholar]
  21. Ornston L. N., Ornston M. K. Glycolate uptake by mutant strains of Escherichia coli K-12. J Bacteriol. 1970 Mar;101(3):1088–1089. doi: 10.1128/jb.101.3.1088-1089.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pouysségur J., Lagarde A. Système de transport du 2-céto-3-désoxy-gluconate chez E. coli K 12: localisation d'un gène de structure et de son opérateur. Mol Gen Genet. 1973 Mar 1;121(2):163–180. doi: 10.1007/BF00277530. [DOI] [PubMed] [Google Scholar]
  23. Rayman M. K., Lo T. C., Sanwal B. D. Transport of succinate in Escherichia coli. II. Characteristics of uptake and energy coupling with transport in membrane preparations. J Biol Chem. 1972 Oct 10;247(19):6332–6339. [PubMed] [Google Scholar]
  24. Saier M. H., Jr, Wentzel D. L., Feucht B. U., Judice J. J. A transport system for phosphoenolpyruvate, 2-phosphoglycerate, and 3-phosphoglycerate in Salmonella typhimurium. J Biol Chem. 1975 Jul 10;250(13):5089–5096. [PubMed] [Google Scholar]
  25. Sweet G. D., Kay C. M., Kay W. W. Tricarboxylate-binding proteins of Salmonella typhimurium. Purification, crystallization, and physical properties. J Biol Chem. 1984 Feb 10;259(3):1586–1592. [PubMed] [Google Scholar]
  26. Wagner C., Odom R., Briggs W. T. The uptake of acetate by Escherichia coli w. Biochem Biophys Res Commun. 1972 Jun 9;47(5):1036–1043. doi: 10.1016/0006-291x(72)90937-0. [DOI] [PubMed] [Google Scholar]
  27. Wilson D. B. Cellular transport mechanisms. Annu Rev Biochem. 1978;47:933–965. doi: 10.1146/annurev.bi.47.070178.004441. [DOI] [PubMed] [Google Scholar]
  28. Winkler H. H., Wilson T. H. The role of energy coupling in the transport of beta-galactosides by Escherichia coli. J Biol Chem. 1966 May 25;241(10):2200–2211. [PubMed] [Google Scholar]

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

RESOURCES