Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1992 May;174(9):2754–2762. doi: 10.1128/jb.174.9.2754-2762.1992

Structure and function of the uhp genes for the sugar phosphate transport system in Escherichia coli and Salmonella typhimurium.

M D Island 1, B Y Wei 1, R J Kadner 1
PMCID: PMC205925  PMID: 1569007

Abstract

Expression of the Escherichia coli sugar phosphate transport system, encoded by the uhpT gene, is regulated by external glucose 6-phosphate through the action of three linked regulatory genes, uhpABC. The nucleotide sequence of the uhp region cloned from Salmonella typhimurium was determined. The deduced Uhp polypeptide sequences from the two organisms are highly related. Comparison with the corrected sequence from E. coli revealed that the four uhp genes are closely spaced, with minimal intergenic distances, and that uhpC is nearly identical in length to uhpT, both of which have substantial sequence relatedness along their entire lengths. To facilitate analysis of uhp gene function, we isolated insertions of a kanamycin resistance (Km) cassette throughout the uhp region. In-frame deletions that removed almost the entire coding region of individual or multiple uhp genes were generated by use of restriction sites at the ends of the Km cassette. The phenotypes of the Km insertions and the in-frame deletions confirmed that all three regulatory genes are required for Uhp function. Whereas the deletion of uhpA completely abolished the expression of a uhpT-lacZ reporter gene, the deletion of uhpB or uhpC resulted in a partially elevated basal level of expression that was not further inducible. These results indicated that UhpB and perhaps UhpC play both positive and negative roles in the control of uhpT transcription. Translational fusions of the uhpBCT genes to topological reporter gene phoA were generated by making use of restriction sites provided by the Km cassette or with transposon TnphoA. The alkaline phosphatase activities of the resultant hybrid proteins were consistent with models predicting that UhpC and UhpT have identical transmembrane topologies, with 10 to 12 transmembrane segments, and that UhpB has 4 to 8 amino-terminal transmembrane segments that anchor the polar carboxyl-terminal half of the protein to the cytoplasmic side of the inner membrane.

Full text

PDF
2754

Selected References

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

  1. Ambudkar S. V., Larson T. J., Maloney P. C. Reconstitution of sugar phosphate transport systems of Escherichia coli. J Biol Chem. 1986 Jul 15;261(20):9083–9086. [PubMed] [Google Scholar]
  2. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Derman A. I., Beckwith J. Escherichia coli alkaline phosphatase fails to acquire disulfide bonds when retained in the cytoplasm. J Bacteriol. 1991 Dec;173(23):7719–7722. doi: 10.1128/jb.173.23.7719-7722.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dietz G. W., Heppel L. A. Studies on the uptake of hexose phosphates. II. The induction of the glucose 6-phosphate transport system by exogenous but not by endogenously formed glucose 6-phosphate. J Biol Chem. 1971 May 10;246(9):2885–2890. [PubMed] [Google Scholar]
  5. Dietz G. W., Jr The hexose phosphate transport system of Escherichia coli. Adv Enzymol Relat Areas Mol Biol. 1976;44:237–259. doi: 10.1002/9780470122891.ch7. [DOI] [PubMed] [Google Scholar]
  6. Ehrmann M., Boyd D., Beckwith J. Genetic analysis of membrane protein topology by a sandwich gene fusion approach. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7574–7578. doi: 10.1073/pnas.87.19.7574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Eidels L., Rick P. D., Stimler N. P., Osborn M. J. Transport of D-arabinose-5-phosphate and D-sedoheptulose-7-phosphate by the hexose phosphate transport system of Salmonella typhimurium. J Bacteriol. 1974 Jul;119(1):138–143. doi: 10.1128/jb.119.1.138-143.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Eiglmeier K., Boos W., Cole S. T. Nucleotide sequence and transcriptional startpoint of the glpT gene of Escherichia coli: extensive sequence homology of the glycerol-3-phosphate transport protein with components of the hexose-6-phosphate transport system. Mol Microbiol. 1987 Nov;1(3):251–258. doi: 10.1111/j.1365-2958.1987.tb01931.x. [DOI] [PubMed] [Google Scholar]
  9. Friedrich M. J., Kadner R. J. Nucleotide sequence of the uhp region of Escherichia coli. J Bacteriol. 1987 Aug;169(8):3556–3563. doi: 10.1128/jb.169.8.3556-3563.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Goldrick D., Yu G. Q., Jiang S. Q., Hong J. S. Nucleotide sequence and transcription start point of the phosphoglycerate transporter gene of Salmonella typhimurium. J Bacteriol. 1988 Aug;170(8):3421–3426. doi: 10.1128/jb.170.8.3421-3426.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Greener A., Hill C. W. Identification of a novel genetic element in Escherichia coli K-12. J Bacteriol. 1980 Oct;144(1):312–321. doi: 10.1128/jb.144.1.312-321.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jones R. N., Wilson H. W., Novick W. J., Jr In vitro evaluation of pyridine-2-azo-p-dimethylaniline cephalosporin, a new diagnostic chromogenic reagent, and comparison with nitrocefin, cephacetrile, and other beta-lactam compounds. J Clin Microbiol. 1982 Apr;15(4):677–683. doi: 10.1128/jcm.15.4.677-683.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kadner R. J., Shattuck-Eidens D. M. Genetic control of the hexose phosphate transport system of Escherichia coli: mapping of deletion and insertion mutations in the uhp region. J Bacteriol. 1983 Sep;155(3):1052–1061. doi: 10.1128/jb.155.3.1052-1061.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kofoid E. C., Parkinson J. S. Transmitter and receiver modules in bacterial signaling proteins. Proc Natl Acad Sci U S A. 1988 Jul;85(14):4981–4985. doi: 10.1073/pnas.85.14.4981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lloyd A. D., Kadner R. J. Topology of the Escherichia coli uhpT sugar-phosphate transporter analyzed by using TnphoA fusions. J Bacteriol. 1990 Apr;172(4):1688–1693. doi: 10.1128/jb.172.4.1688-1693.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Manoil C., Beckwith J. TnphoA: a transposon probe for protein export signals. Proc Natl Acad Sci U S A. 1985 Dec;82(23):8129–8133. doi: 10.1073/pnas.82.23.8129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Merkel T. J., Nelson D. M., Brauer C. L., Kadner R. J. Promoter elements required for positive control of transcription of the Escherichia coli uhpT gene. J Bacteriol. 1992 May;174(9):2763–2770. doi: 10.1128/jb.174.9.2763-2770.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Michaelis S., Inouye H., Oliver D., Beckwith J. Mutations that alter the signal sequence of alkaline phosphatase in Escherichia coli. J Bacteriol. 1983 Apr;154(1):366–374. doi: 10.1128/jb.154.1.366-374.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Oishi M., Cosloy S. D. The genetic and biochemical basis of the transformability of Escherichia coli K12. Biochem Biophys Res Commun. 1972 Dec 18;49(6):1568–1572. doi: 10.1016/0006-291x(72)90520-7. [DOI] [PubMed] [Google Scholar]
  20. Pogell B. M., Maity B. R., Frumkin S., Shapiro S. Induction of an active transport system for glucose 6-phosphate in Escherichia coli. Arch Biochem Biophys. 1966 Sep 26;116(1):406–415. doi: 10.1016/0003-9861(66)90047-6. [DOI] [PubMed] [Google Scholar]
  21. Roland K. L., Liu C. G., Turnbough C. L., Jr Role of the ribosome in suppressing transcriptional termination at the pyrBI attenuator of Escherichia coli K-12. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7149–7153. doi: 10.1073/pnas.85.19.7149. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Shattuck-Eidens D. M., Kadner R. J. Exogenous induction of the Escherichia coli hexose phosphate transport system defined by uhp-lac operon fusions. J Bacteriol. 1981 Oct;148(1):203–209. doi: 10.1128/jb.148.1.203-209.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Simons R. W., Houman F., Kleckner N. Improved single and multicopy lac-based cloning vectors for protein and operon fusions. Gene. 1987;53(1):85–96. doi: 10.1016/0378-1119(87)90095-3. [DOI] [PubMed] [Google Scholar]
  24. Sonna L. A., Ambudkar S. V., Maloney P. C. The mechanism of glucose 6-phosphate transport by Escherichia coli. J Biol Chem. 1988 May 15;263(14):6625–6630. [PubMed] [Google Scholar]
  25. Sorge J. A., Blinderman L. A. ExoMeth sequencing of DNA: eliminating the need for subcloning and oligonucleotide primers. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9208–9212. doi: 10.1073/pnas.86.23.9208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Stock J. B., Ninfa A. J., Stock A. M. Protein phosphorylation and regulation of adaptive responses in bacteria. Microbiol Rev. 1989 Dec;53(4):450–490. doi: 10.1128/mr.53.4.450-490.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Weston L. A., Kadner R. J. Identification of uhp polypeptides and evidence for their role in exogenous induction of the sugar phosphate transport system of Escherichia coli K-12. J Bacteriol. 1987 Aug;169(8):3546–3555. doi: 10.1128/jb.169.8.3546-3555.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Weston L. A., Kadner R. J. Role of uhp genes in expression of the Escherichia coli sugar-phosphate transport system. J Bacteriol. 1988 Aug;170(8):3375–3383. doi: 10.1128/jb.170.8.3375-3383.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Winans S. C., Elledge S. J., Krueger J. H., Walker G. C. Site-directed insertion and deletion mutagenesis with cloned fragments in Escherichia coli. J Bacteriol. 1985 Mar;161(3):1219–1221. doi: 10.1128/jb.161.3.1219-1221.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Winkler H. H. Compartmentation in the induction of the hexose-6-phosphate transport system of Escherichia coli. J Bacteriol. 1970 Feb;101(2):470–475. doi: 10.1128/jb.101.2.470-475.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES