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. 1989 Sep;86(18):6953–6957. doi: 10.1073/pnas.86.18.6953

Reconstitution of a bacterial periplasmic permease in proteoliposomes and demonstration of ATP hydrolysis concomitant with transport.

L Bishop 1, R Agbayani Jr 1, S V Ambudkar 1, P C Maloney 1, G F Ames 1
PMCID: PMC297969  PMID: 2674940

Abstract

The histidine periplasmic permease of Salmonella typhimurium has been partially purified and reconstituted into proteoliposomes. In this in vitro preparation, transport activity is completely dependent on the presence of all four permease proteins (HisJ, HisQ, HisM, and HisP) and on internal ATP. The reconstituted system shows initial rates of transport that are comparable to those obtained with right-side-out membrane vesicles and it establishes a 100-fold concentration gradient for histidine. Proteoliposomes also transport histidine when GTP replaces ATP. Proteoliposomes do not catalyze significant ATP hydrolysis until histidine transport is initiated by addition of substrate along with HisJ, the water-soluble histidine-binding protein. Both initially and throughout the course of substrate transport there is a concomitant hydrolysis of ATP, with an apparent stoichiometry (ATP/histidine) of 5:1. These experiments demonstrate directly that ATP is the source of energy for periplasmic permeases, thus resolving previous controversies on this topic.

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Selected References

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

  1. Ambudkar S. V., Maloney P. C. Bacterial anion exchange. Use of osmolytes during solubilization and reconstitution of phosphate-linked antiport from Streptococcus lactis. J Biol Chem. 1986 Aug 5;261(22):10079–10086. [PubMed] [Google Scholar]
  2. Ames G. F. Bacterial periplasmic transport systems: structure, mechanism, and evolution. Annu Rev Biochem. 1986;55:397–425. doi: 10.1146/annurev.bi.55.070186.002145. [DOI] [PubMed] [Google Scholar]
  3. Ames G. F., Nikaido K., Groarke J., Petithory J. Reconstitution of periplasmic transport in inside-out membrane vesicles. Energization by ATP. J Biol Chem. 1989 Mar 5;264(7):3998–4002. [PubMed] [Google Scholar]
  4. Bell A. W., Buckel S. D., Groarke J. M., Hope J. N., Kingsley D. H., Hermodson M. A. The nucleotide sequences of the rbsD, rbsA, and rbsC genes of Escherichia coli K12. J Biol Chem. 1986 Jun 15;261(17):7652–7658. [PubMed] [Google Scholar]
  5. Berger E. A., Heppel L. A. Different mechanisms of energy coupling for the shock-sensitive and shock-resistant amino acid permeases of Escherichia coli. J Biol Chem. 1974 Dec 25;249(24):7747–7755. [PubMed] [Google Scholar]
  6. Doolittle R. F., Johnson M. S., Husain I., Van Houten B., Thomas D. C., Sancar A. Domainal evolution of a prokaryotic DNA repair protein and its relationship to active-transport proteins. Nature. 1986 Oct 2;323(6087):451–453. doi: 10.1038/323451a0. [DOI] [PubMed] [Google Scholar]
  7. Ferro-Luzzi Ames G. The basis of multidrug resistance in mammalian cells: homology with bacterial transport. Cell. 1986 Nov 7;47(3):323–324. doi: 10.1016/0092-8674(86)90585-4. [DOI] [PubMed] [Google Scholar]
  8. Gallagher M. P., Pearce S. R., Higgins C. F. Identification and localization of the membrane-associated, ATP-binding subunit of the oligopeptide permease of Salmonella typhimurium. Eur J Biochem. 1989 Mar 1;180(1):133–141. doi: 10.1111/j.1432-1033.1989.tb14623.x. [DOI] [PubMed] [Google Scholar]
  9. Higgins C. F., Hiles I. D., Salmond G. P., Gill D. R., Downie J. A., Evans I. J., Holland I. B., Gray L., Buckel S. D., Bell A. W. A family of related ATP-binding subunits coupled to many distinct biological processes in bacteria. Nature. 1986 Oct 2;323(6087):448–450. doi: 10.1038/323448a0. [DOI] [PubMed] [Google Scholar]
  10. Higgins C. F., Hiles I. D., Whalley K., Jamieson D. J. Nucleotide binding by membrane components of bacterial periplasmic binding protein-dependent transport systems. EMBO J. 1985 Apr;4(4):1033–1039. doi: 10.1002/j.1460-2075.1985.tb03735.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hobson A. C., Weatherwax R., Ames G. F. ATP-binding sites in the membrane components of histidine permease, a periplasmic transport system. Proc Natl Acad Sci U S A. 1984 Dec;81(23):7333–7337. doi: 10.1073/pnas.81.23.7333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hunt A. G., Hong J. The reconstitution of binding protein-dependent active transport of glutamine in isolated membrane vesicles from Escherichia coli. J Biol Chem. 1981 Dec 10;256(23):11988–11991. [PubMed] [Google Scholar]
  13. Joshi A. K., Ahmed S., Ferro-Luzzi Ames G. Energy coupling in bacterial periplasmic transport systems. Studies in intact Escherichia coli cells. J Biol Chem. 1989 Feb 5;264(4):2126–2133. [PubMed] [Google Scholar]
  14. Kaback H. R. The lac carrier protein in Escherichia coli. J Membr Biol. 1983;76(2):95–112. doi: 10.1007/BF02000610. [DOI] [PubMed] [Google Scholar]
  15. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  16. Maloney P. C., Ambudkar S. V. Functional reconstitution of prokaryote and eukaryote membrane proteins. Arch Biochem Biophys. 1989 Feb 15;269(1):1–10. doi: 10.1016/0003-9861(89)90080-5. [DOI] [PubMed] [Google Scholar]
  17. Maloney P. C. Energy coupling to ATP synthesis by the proton-translocating ATPase. J Membr Biol. 1982;67(1):1–12. doi: 10.1007/BF01868643. [DOI] [PubMed] [Google Scholar]
  18. Mount S. M. Sequence similarity. Nature. 1987 Feb 5;325(6104):487–487. doi: 10.1038/325487c0. [DOI] [PubMed] [Google Scholar]
  19. Newman M. J., Foster D. L., Wilson T. H., Kaback H. R. Purification and reconstitution of functional lactose carrier from Escherichia coli. J Biol Chem. 1981 Nov 25;256(22):11804–11808. [PubMed] [Google Scholar]
  20. Newman M. J., Wilson T. H. Solubilization and reconstitution of the lactose transport system from Escherichia coli. J Biol Chem. 1980 Nov 25;255(22):10583–10586. [PubMed] [Google Scholar]
  21. Peterson G. L. A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal Biochem. 1977 Dec;83(2):346–356. doi: 10.1016/0003-2697(77)90043-4. [DOI] [PubMed] [Google Scholar]
  22. Prossnitz E., Gee A., Ames G. F. Reconstitution of the histidine periplasmic transport system in membrane vesicles. Energy coupling and interaction between the binding protein and the membrane complex. J Biol Chem. 1989 Mar 25;264(9):5006–5014. [PubMed] [Google Scholar]
  23. Smith G. S., Scholes P. B. The H+/ATP stoichiometry of the (H+ +K+)-ATPase of dog gastric microsomes. Biochim Biophys Acta. 1982 Jun 28;688(3):803–807. doi: 10.1016/0005-2736(82)90295-4. [DOI] [PubMed] [Google Scholar]
  24. Walker J. E., Saraste M., Runswick M. J., Gay N. J. Distantly related sequences in the alpha- and beta-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold. EMBO J. 1982;1(8):945–951. doi: 10.1002/j.1460-2075.1982.tb01276.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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