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. 1993 Apr;2(4):506–521. doi: 10.1002/pro.5560020403

Sequence analyses and evolutionary relationships among the energy-coupling proteins Enzyme I and HPr of the bacterial phosphoenolpyruvate: sugar phosphotransferase system.

J Reizer 1, C Hoischen 1, A Reizer 1, T N Pham 1, M H Saier Jr 1
PMCID: PMC2142364  PMID: 7686067

Abstract

We have previously reported the overexpression, purification, and biochemical properties of the Bacillus subtilis Enzyme I of the phosphoenolpyruvate: sugar phosphotransferase system (PTS) (Reizer, J., et al., 1992, J. Biol. Chem. 267, 9158-9169). We now report the sequencing of the ptsI gene of B. subtilis encoding Enzyme I (570 amino acids and 63,076 Da). Putative transcriptional regulatory signals are identified, and the pts operon is shown to be subject to carbon source-dependent regulation. Multiple alignments of the B. subtilis Enzyme I with (1) six other sequenced Enzymes I of the PTS from various bacterial species, (2) phosphoenolpyruvate synthase of Escherichia coli, and (3) bacterial and plant pyruvate: phosphate dikinases (PPDKs) revealed regions of sequence similarity as well as divergence. Statistical analyses revealed that these three types of proteins comprise a homologous family, and the phylogenetic tree of the 11 sequenced protein members of this family was constructed. This tree was compared with that of the 12 sequence HPr proteins or protein domains. Antibodies raised against the B. subtilis and E. coli Enzymes I exhibited immunological cross-reactivity with each other as well as with PPDK of Bacteroides symbiosus, providing support for the evolutionary relationships of these proteins suggested from the sequence comparisons. Putative flexible linkers tethering the N-terminal and the C-terminal domains of protein members of the Enzyme I family were identified, and their potential significance with regard to Enzyme I function is discussed. The codon choice pattern of the B. subtilis and E. coli ptsI and ptsH genes was found to exhibit a bias toward optimal codons in these organisms.(ABSTRACT TRUNCATED AT 250 WORDS)

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Selected References

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  1. Bairoch A. PROSITE: a dictionary of sites and patterns in proteins. Nucleic Acids Res. 1992 May 11;20 (Suppl):2013–2018. doi: 10.1093/nar/20.suppl.2013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brennan R. G., Matthews B. W. The helix-turn-helix DNA binding motif. J Biol Chem. 1989 Feb 5;264(4):1903–1906. [PubMed] [Google Scholar]
  3. Burnell J. N., Hatch M. D. Activation and inactivation of an enzyme catalyzed by a single, bifunctional protein: a new example and why. Arch Biochem Biophys. 1986 Mar;245(2):297–304. doi: 10.1016/0003-9861(86)90219-5. [DOI] [PubMed] [Google Scholar]
  4. Chen Y., Reizer J., Saier M. H., Jr, Fairbrother W. J., Wright P. E. Mapping of the binding interfaces of the proteins of the bacterial phosphotransferase system, HPr and IIAglc. Biochemistry. 1993 Jan 12;32(1):32–37. doi: 10.1021/bi00052a006. [DOI] [PubMed] [Google Scholar]
  5. Cohen P. The role of cyclic-AMP-dependent protein kinase in the regulation of glycogen metabolism in mammalian skeletal muscle. Curr Top Cell Regul. 1978;14:117–196. doi: 10.1016/b978-0-12-152814-0.50008-3. [DOI] [PubMed] [Google Scholar]
  6. Dayhoff M. O., Barker W. C., Hunt L. T. Establishing homologies in protein sequences. Methods Enzymol. 1983;91:524–545. doi: 10.1016/s0076-6879(83)91049-2. [DOI] [PubMed] [Google Scholar]
  7. De Reuse H., Danchin A. Positive regulation of the pts operon of Escherichia coli: genetic evidence for a signal transduction mechanism. J Bacteriol. 1991 Jan;173(2):727–733. doi: 10.1128/jb.173.2.727-733.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. De Reuse H., Danchin A. The ptsH, ptsI, and crr genes of the Escherichia coli phosphoenolpyruvate-dependent phosphotransferase system: a complex operon with several modes of transcription. J Bacteriol. 1988 Sep;170(9):3827–3837. doi: 10.1128/jb.170.9.3827-3837.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. De Reuse H., Roy A., Danchin A. Analysis of the ptsH-ptsI-crr region in Escherichia coli K-12: nucleotide sequence of the ptsH gene. Gene. 1985;35(1-2):199–207. doi: 10.1016/0378-1119(85)90172-6. [DOI] [PubMed] [Google Scholar]
  10. Deutscher J., Pevec B., Beyreuther K., Kiltz H. H., Hengstenberg W. Streptococcal phosphoenolpyruvate-sugar phosphotransferase system: amino acid sequence and site of ATP-dependent phosphorylation of HPr. Biochemistry. 1986 Oct 21;25(21):6543–6551. doi: 10.1021/bi00369a031. [DOI] [PubMed] [Google Scholar]
  11. Eisermann R., Fischer R., Kessler U., Neubauer A., Hengstenberg W. Staphylococcal phosphoenolpyruvate-dependent phosphotransferase system. Purification and protein sequencing of the Staphylococcus carnosus histidine-containing protein, and cloning and DNA sequencing of the ptsH gene. Eur J Biochem. 1991 Apr 10;197(1):9–14. doi: 10.1111/j.1432-1033.1991.tb15875.x. [DOI] [PubMed] [Google Scholar]
  12. Fairbrother W. J., Cavanagh J., Dyson H. J., Palmer A. G., 3rd, Sutrina S. L., Reizer J., Saier M. H., Jr, Wright P. E. Polypeptide backbone resonance assignments and secondary structure of Bacillus subtilis enzyme IIIglc determined by two-dimensional and three-dimensional heteronuclear NMR spectroscopy. Biochemistry. 1991 Jul 16;30(28):6896–6907. doi: 10.1021/bi00242a013. [DOI] [PubMed] [Google Scholar]
  13. Fairbrother W. J., Gippert G. P., Reizer J., Saier M. H., Jr, Wright P. E. Low resolution solution structure of the Bacillus subtilis glucose permease IIA domain derived from heteronuclear three-dimensional NMR spectroscopy. FEBS Lett. 1992 Jan 20;296(2):148–152. doi: 10.1016/0014-5793(92)80367-p. [DOI] [PubMed] [Google Scholar]
  14. Feng D. F., Doolittle R. F. Progressive alignment and phylogenetic tree construction of protein sequences. Methods Enzymol. 1990;183:375–387. doi: 10.1016/0076-6879(90)83025-5. [DOI] [PubMed] [Google Scholar]
  15. Gonzy-Tréboul G., Steinmetz M. Phosphoenolpyruvate:sugar phosphotransferase system of Bacillus subtilis: cloning of the region containing the ptsH and ptsI genes and evidence for a crr-like gene. J Bacteriol. 1987 May;169(5):2287–2290. doi: 10.1128/jb.169.5.2287-2290.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gonzy-Tréboul G., Zagorec M., Rain-Guion M. C., Steinmetz M. Phosphoenolpyruvate:sugar phosphotransferase system of Bacillus subtilis: nucleotide sequence of ptsX, ptsH and the 5'-end of ptsI and evidence for a ptsHI operon. Mol Microbiol. 1989 Jan;3(1):103–112. doi: 10.1111/j.1365-2958.1989.tb00109.x. [DOI] [PubMed] [Google Scholar]
  17. Goss N. H., Evans C. T., Wood H. G. Pyruvate phosphate dikinase: sequence of the histidyl peptide, the pyrophosphoryl and phosphoryl carrier. Biochemistry. 1980 Dec 9;19(25):5805–5809. doi: 10.1021/bi00566a022. [DOI] [PubMed] [Google Scholar]
  18. Gouy M., Gautier C. Codon usage in bacteria: correlation with gene expressivity. Nucleic Acids Res. 1982 Nov 25;10(22):7055–7074. doi: 10.1093/nar/10.22.7055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Herzberg O., Reddy P., Sutrina S., Saier M. H., Jr, Reizer J., Kapadia G. Structure of the histidine-containing phosphocarrier protein HPr from Bacillus subtilis at 2.0-A resolution. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2499–2503. doi: 10.1073/pnas.89.6.2499. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ikemura T. Codon usage and tRNA content in unicellular and multicellular organisms. Mol Biol Evol. 1985 Jan;2(1):13–34. doi: 10.1093/oxfordjournals.molbev.a040335. [DOI] [PubMed] [Google Scholar]
  21. Jenkinson H. F. Properties of a phosphocarrier protein (HPr) extracted from intact cells of Streptococcus sanguis. J Gen Microbiol. 1989 Dec;135(12):3183–3197. doi: 10.1099/00221287-135-12-3183. [DOI] [PubMed] [Google Scholar]
  22. Kohlbrecher D., Eisermann R., Hengstenberg W. Staphylococcal phosphoenolpyruvate-dependent phosphotransferase system: molecular cloning and nucleotide sequence of the Staphylococcus carnosus ptsI gene and expression and complementation studies of the gene product. J Bacteriol. 1992 Apr;174(7):2208–2214. doi: 10.1128/jb.174.7.2208-2214.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kumar A., Larsen C. E., Preiss J. Biosynthesis of bacterial glycogen. Primary structure of Escherichia coli ADP-glucose:alpha-1,4-glucan, 4-glucosyltransferase as deduced from the nucleotide sequence of the glgA gene. J Biol Chem. 1986 Dec 5;261(34):16256–16259. [PubMed] [Google Scholar]
  24. LiCalsi C., Crocenzi T. S., Freire E., Roseman S. Sugar transport by the bacterial phosphotransferase system. Structural and thermodynamic domains of enzyme I of Salmonella typhimurium. J Biol Chem. 1991 Oct 15;266(29):19519–19527. [PubMed] [Google Scholar]
  25. Liao D. I., Kapadia G., Reddy P., Saier M. H., Jr, Reizer J., Herzberg O. Structure of the IIA domain of the glucose permease of Bacillus subtilis at 2.2-A resolution. Biochemistry. 1991 Oct 8;30(40):9583–9594. doi: 10.1021/bi00104a004. [DOI] [PubMed] [Google Scholar]
  26. Mattoo R. L., Waygood E. B. Determination of the levels of HPr and enzyme I of the phosphoenolpyruvate-sugar phosphotransferase system in Escherichia coli and Salmonella typhimurium. Can J Biochem Cell Biol. 1983 Jan;61(1):29–37. doi: 10.1139/o83-005. [DOI] [PubMed] [Google Scholar]
  27. Meadow N. D., Fox D. K., Roseman S. The bacterial phosphoenolpyruvate: glycose phosphotransferase system. Annu Rev Biochem. 1990;59:497–542. doi: 10.1146/annurev.bi.59.070190.002433. [DOI] [PubMed] [Google Scholar]
  28. Narindrasorasak S., Bridger W. A. Phosphoenolypyruvate synthetase of Escherichia coli: molecular weight, subunit composition, and identification of phosphohistidine in phosphoenzyme intermediate. J Biol Chem. 1977 May 25;252(10):3121–3127. [PubMed] [Google Scholar]
  29. Orchard L. M., Kornberg H. L. Sequence similarities between the gene specifying 1-phosphofructokinase (fruK), genes specifying other kinases in Escherichia coli K12, and lacC of Staphylococcus aureus. Proc Biol Sci. 1990 Nov 22;242(1304):87–90. doi: 10.1098/rspb.1990.0108. [DOI] [PubMed] [Google Scholar]
  30. Pearson W. R., Lipman D. J. Improved tools for biological sequence comparison. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444–2448. doi: 10.1073/pnas.85.8.2444. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Platt T. Termination of transcription and its regulation in the tryptophan operon of E. coli. Cell. 1981 Apr;24(1):10–23. doi: 10.1016/0092-8674(81)90496-7. [DOI] [PubMed] [Google Scholar]
  32. Pocalyko D. J., Carroll L. J., Martin B. M., Babbitt P. C., Dunaway-Mariano D. Analysis of sequence homologies in plant and bacterial pyruvate phosphate dikinase, enzyme I of the bacterial phosphoenolpyruvate: sugar phosphotransferase system and other PEP-utilizing enzymes. Identification of potential catalytic and regulatory motifs. Biochemistry. 1990 Dec 4;29(48):10757–10765. doi: 10.1021/bi00500a006. [DOI] [PubMed] [Google Scholar]
  33. Reizer J., Deutscher J., Saier M. H., Jr Metabolite-sensitive, ATP-dependent, protein kinase-catalyzed phosphorylation of HPr, a phosphocarrier protein of the phosphotransferase system in gram-positive bacteria. Biochimie. 1989 Sep-Oct;71(9-10):989–996. doi: 10.1016/0300-9084(89)90102-8. [DOI] [PubMed] [Google Scholar]
  34. Reizer J., Novotny M. J., Hengstenberg W., Saier M. H., Jr Properties of ATP-dependent protein kinase from Streptococcus pyogenes that phosphorylates a seryl residue in HPr, a phosphocarrier protein of the phosphotransferase system. J Bacteriol. 1984 Oct;160(1):333–340. doi: 10.1128/jb.160.1.333-340.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Reizer J., Peterkofsky A., Romano A. H. Evidence for the presence of heat-stable protein (HPr) and ATP-dependent HPr kinase in heterofermentative lactobacilli lacking phosphoenolpyruvate:glycose phosphotransferase activity. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2041–2045. doi: 10.1073/pnas.85.7.2041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Reizer J. Regulation of sugar uptake and efflux in gram-positive bacteria. FEMS Microbiol Rev. 1989 Jun;5(1-2):149–156. doi: 10.1016/0168-6445(89)90019-3. [DOI] [PubMed] [Google Scholar]
  37. Reizer J., Saier M. H., Jr, Deutscher J., Grenier F., Thompson J., Hengstenberg W. The phosphoenolpyruvate:sugar phosphotransferase system in gram-positive bacteria: properties, mechanism, and regulation. Crit Rev Microbiol. 1988;15(4):297–338. doi: 10.3109/10408418809104461. [DOI] [PubMed] [Google Scholar]
  38. Reizer J., Sutrina S. L., Saier M. H., Stewart G. C., Peterkofsky A., Reddy P. Mechanistic and physiological consequences of HPr(ser) phosphorylation on the activities of the phosphoenolpyruvate:sugar phosphotransferase system in gram-positive bacteria: studies with site-specific mutants of HPr. EMBO J. 1989 Jul;8(7):2111–2120. doi: 10.1002/j.1460-2075.1989.tb03620.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Reizer J., Sutrina S. L., Wu L. F., Deutscher J., Reddy P., Saier M. H., Jr Functional interactions between proteins of the phosphoenolpyruvate:sugar phosphotransferase systems of Bacillus subtilis and Escherichia coli. J Biol Chem. 1992 May 5;267(13):9158–9169. [PubMed] [Google Scholar]
  40. Rephaeli A. W., Saier M. H., Jr Regulation of genes coding for enzyme constituents of the bacterial phosphotransferase system. J Bacteriol. 1980 Feb;141(2):658–663. doi: 10.1128/jb.141.2.658-663.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rosche E., Westhoff P. Primary structure of pyruvate, orthophosphate dikinase in the dicotyledonous C4 plant Flaveria trinervia. FEBS Lett. 1990 Oct 29;273(1-2):116–121. doi: 10.1016/0014-5793(90)81064-u. [DOI] [PubMed] [Google Scholar]
  42. Saffen D. W., Presper K. A., Doering T. L., Roseman S. Sugar transport by the bacterial phosphotransferase system. Molecular cloning and structural analysis of the Escherichia coli ptsH, ptsI, and crr genes. J Biol Chem. 1987 Nov 25;262(33):16241–16253. [PubMed] [Google Scholar]
  43. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Sharp P. M., Devine K. M. Codon usage and gene expression level in Dictyostelium discoideum: highly expressed genes do 'prefer' optimal codons. Nucleic Acids Res. 1989 Jul 11;17(13):5029–5039. doi: 10.1093/nar/17.13.5029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Sharp P. M., Li W. H. An evolutionary perspective on synonymous codon usage in unicellular organisms. J Mol Evol. 1986;24(1-2):28–38. doi: 10.1007/BF02099948. [DOI] [PubMed] [Google Scholar]
  46. Sharp P. M., Li W. H. The codon Adaptation Index--a measure of directional synonymous codon usage bias, and its potential applications. Nucleic Acids Res. 1987 Feb 11;15(3):1281–1295. doi: 10.1093/nar/15.3.1281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Shields D. C., Sharp P. M. Synonymous codon usage in Bacillus subtilis reflects both translational selection and mutational biases. Nucleic Acids Res. 1987 Oct 12;15(19):8023–8040. doi: 10.1093/nar/15.19.8023. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Soberon X., Covarrubias L., Bolivar F. Construction and characterization of new cloning vehicles. IV. Deletion derivatives of pBR322 and pBR325. Gene. 1980 May;9(3-4):287–305. doi: 10.1016/0378-1119(90)90328-o. [DOI] [PubMed] [Google Scholar]
  49. Stone M. J., Fairbrother W. J., Palmer A. G., 3rd, Reizer J., Saier M. H., Jr, Wright P. E. Backbone dynamics of the Bacillus subtilis glucose permease IIA domain determined from 15N NMR relaxation measurements. Biochemistry. 1992 May 12;31(18):4394–4406. doi: 10.1021/bi00133a003. [DOI] [PubMed] [Google Scholar]
  50. Thibault L., Vadeboncoeur C. Phosphoenolpyruvate-sugar phosphotransferase transport system of Streptococcus mutans: purification of HPr and enzyme I and determination of their intracellular concentrations by rocket immunoelectrophoresis. Infect Immun. 1985 Dec;50(3):817–825. doi: 10.1128/iai.50.3.817-825.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wittekind M., Reizer J., Deutscher J., Saier M. H., Klevit R. E. Common structural changes accompany the functional inactivation of HPr by seryl phosphorylation or by serine to aspartate substitution. Biochemistry. 1989 Dec 26;28(26):9908–9912. doi: 10.1021/bi00452a005. [DOI] [PubMed] [Google Scholar]
  52. Wittekind M., Reizer J., Klevit R. E. Sequence-specific 1H NMR resonance assignments of Bacillus subtilis HPr: use of spectra obtained from mutants to resolve spectral overlap. Biochemistry. 1990 Aug 7;29(31):7191–7200. doi: 10.1021/bi00483a006. [DOI] [PubMed] [Google Scholar]
  53. Wood H. G., O'brien W. E., Micheales G. Properties of carboxytransphosphorylase; pyruvate, phosphate dikinase; pyrophosphate-phosphofructikinase and pyrophosphate-acetate kinase and their roles in the metabolism of inorganic pyrophosphate. Adv Enzymol Relat Areas Mol Biol. 1977;45:85–155. doi: 10.1002/9780470122907.ch2. [DOI] [PubMed] [Google Scholar]
  54. Wu L. F., Tomich J. M., Saier M. H., Jr Structure and evolution of a multidomain multiphosphoryl transfer protein. Nucleotide sequence of the fruB(HI) gene in Rhodobacter capsulatus and comparisons with homologous genes from other organisms. J Mol Biol. 1990 Jun 20;213(4):687–703. doi: 10.1016/S0022-2836(05)80256-6. [DOI] [PubMed] [Google Scholar]

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