1887

Abstract

In the extremely thermophilic bacterium HB8, one of the four TetR-family transcriptional regulators, which we named . FadR, negatively regulated the expression of several genes, including those involved in fatty acid degradation, both and . . FadR repressed the expression of the target genes by binding pseudopalindromic sequences covering the predicted −10 hexamers of their promoters, and medium-to-long straight-chain (C10–18) fatty acyl-CoA molecules were effective for transcriptional derepression. An X-ray crystal structure analysis revealed that . FadR bound one lauroyl (C12)-CoA molecule per FadR monomer, with its acyl chain moiety in the centre of the FadR molecule, enclosed within a tunnel-like substrate-binding pocket surrounded by hydrophobic residues, and the CoA moiety interacting with basic residues on the protein surface. The growth of . HB8, with palmitic acid as the sole carbon source, increased the expression of FadR-regulated genes. These results indicate that in . HB8, medium-to-long straight-chain fatty acids can be used for metabolic energy under the control of FadR, although the major fatty acids found in this strain are - and -branched-chain (C15 and 17) fatty acids.

Funding
This study was supported by the:
  • Ministry of Education, Culture, Sports, Science and Technology, Japan (Award 22510208)
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.048017-0
2011-06-01
2021-08-05
Loading full text...

Full text loading...

/deliver/fulltext/micro/157/6/1589.html?itemId=/content/journal/micro/10.1099/mic.0.048017-0&mimeType=html&fmt=ahah

References

  1. Agari Y., Kashihara A., Yokoyama S., Kuramitsu S., Shinkai A. ( 2008). Global gene expression mediated by Thermus thermophilus SdrP, a CRP/FNR family transcriptional regulator. Mol Microbiol 70:60–75 [View Article][PubMed]
    [Google Scholar]
  2. Baker N. A., Sept D., Joseph S., Holst M. J., McCammon J. A. ( 2001). Electrostatics of nanosystems: application to microtubules and the ribosome. Proc Natl Acad Sci U S A 98:10037–10041 [View Article][PubMed]
    [Google Scholar]
  3. Brünger A. T., Adams P. D., Clore G. M., DeLano W. L., Gros P., Grosse-Kunstleve R. W., Jiang J. S., Kuszewski J., Nilges M. et al. ( 1998). Crystallography & NMR System: a new software suite for macromolecular structure determination. Acta Crystallogr D Biol Crystallogr 54:905–921 [View Article][PubMed]
    [Google Scholar]
  4. Campbell J. W., Cronan J. E. Jr ( 2001). Escherichia coli FadR positively regulates transcription of the fabB fatty acid biosynthetic gene. J Bacteriol 183:5982–5990 [View Article][PubMed]
    [Google Scholar]
  5. Cho B. K., Knight E. M., Palsson B. O. ( 2006). Transcriptional regulation of the fad regulon genes of Escherichia coli by ArcA. Microbiology 152:2207–2219 [View Article][PubMed]
    [Google Scholar]
  6. Cronan J. E. Jr, Subrahmanyam S. ( 1998). FadR, transcriptional co-ordination of metabolic expediency. Mol Microbiol 29:937–943 [View Article][PubMed]
    [Google Scholar]
  7. Crooks G. E., Hon G., Chandonia J. M., Brenner S. E. ( 2004). WebLogo: a sequence logo generator. Genome Res 14:1188–1190 [View Article][PubMed]
    [Google Scholar]
  8. DiRusso C. C., Nyström T. ( 1998). The fats of Escherichia coli during infancy and old age: regulation by global regulators, alarmones and lipid intermediates. Mol Microbiol 27:1–8 [View Article][PubMed]
    [Google Scholar]
  9. DiRusso C. C., Heimert T. L., Metzger A. K. ( 1992). Characterization of FadR, a global transcriptional regulator of fatty acid metabolism in Escherichia coli. Interaction with the fadB promoter is prevented by long chain fatty acyl coenzyme A. J Biol Chem 267:8685–8691[PubMed]
    [Google Scholar]
  10. Emsley P., Cowtan K. ( 2004). Coot: model-building tools for molecular graphics. Acta Crystallogr D Biol Crystallogr 60:2126–2132 [View Article][PubMed]
    [Google Scholar]
  11. Feng Y., Cronan J. E. ( 2010). Overlapping repressor binding sites result in additive regulation of Escherichia coli FadH by FadR and ArcA. J Bacteriol 192:4289–4299 [View Article][PubMed]
    [Google Scholar]
  12. Fujita Y., Matsuoka H., Hirooka K. ( 2007). Regulation of fatty acid metabolism in bacteria. Mol Microbiol 66:829–839 [View Article][PubMed]
    [Google Scholar]
  13. Gouet P., Robert X., Courcelle E. ( 2003). ESPript/ENDscript: extracting and rendering sequence and 3D information from atomic structures of proteins. Nucleic Acids Res 31:3320–3323 [View Article][PubMed]
    [Google Scholar]
  14. Grkovic S., Brown M. H., Roberts N. J., Paulsen I. T., Skurray R. A. ( 1998). QacR is a repressor protein that regulates expression of the Staphylococcus aureus multidrug efflux pump QacA. J Biol Chem 273:18665–18673 [View Article][PubMed]
    [Google Scholar]
  15. Hashimoto Y., Yano T., Kuramitsu S., Kagamiyama H. ( 2001). Disruption of Thermus thermophilus genes by homologous recombination using a thermostable kanamycin-resistant marker. FEBS Lett 506:231–234 [View Article][PubMed]
    [Google Scholar]
  16. Haydon D. J., Guest J. R. ( 1991). A new family of bacterial regulatory proteins. FEMS Microbiol Lett 63:291–295 [View Article][PubMed]
    [Google Scholar]
  17. Henry M. F., Cronan J. E. Jr ( 1991). Escherichia coli transcription factor that both activates fatty acid synthesis and represses fatty acid degradation. J Mol Biol 222:843–849 [View Article][PubMed]
    [Google Scholar]
  18. Henry M. F., Cronan J. E. Jr ( 1992). A new mechanism of transcriptional regulation: release of an activator triggered by small molecule binding. Cell 70:671–679 [View Article][PubMed]
    [Google Scholar]
  19. Higashitani A., Nishimura Y., Hara H., Aiba H., Mizuno T., Horiuchi K. ( 1993). Osmoregulation of the fatty acid receptor gene fadL in Escherichia coli . Mol Gen Genet 240:339–347[PubMed]
    [Google Scholar]
  20. Hisanaga Y., Ago H., Nakagawa N., Hamada K., Ida K., Yamamoto M., Hori T., Arii Y., Sugahara M. et al. ( 2004). Structural basis of the substrate-specific two-step catalysis of long chain fatty acyl-CoA synthetase dimer. J Biol Chem 279:31717–31726 [View Article][PubMed]
    [Google Scholar]
  21. Holm L., Sander C. ( 1998). Touring protein fold space with Dali/FSSP. Nucleic Acids Res 26:316–319 [View Article][PubMed]
    [Google Scholar]
  22. Kabsch W., Sander C. ( 1983). Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers 22:2577–2637 [View Article][PubMed]
    [Google Scholar]
  23. Kuramitsu S., Hiromi K., Hayashi H., Morino Y., Kagamiyama H. ( 1990). Pre-steady-state kinetics of Escherichia coli aspartate aminotransferase catalyzed reactions and thermodynamic aspects of its substrate specificity. Biochemistry 29:5469–5476 [View Article][PubMed]
    [Google Scholar]
  24. Larkin M. A., Blackshields G., Brown N. P., Chenna R., McGettigan P. A., McWilliam H., Valentin F., Wallace I. M., Wilm A. et al. ( 2007). clustal w and clustal x version 2.0. Bioinformatics 23:2947–2948 [View Article][PubMed]
    [Google Scholar]
  25. LeMaster D. M., Richards F. M. ( 1985). 1H-15N heteronuclear NMR studies of Escherichia coli thioredoxin in samples isotopically labeled by residue type. Biochemistry 24:7263–7268 [View Article][PubMed]
    [Google Scholar]
  26. Marchler-Bauer A., Panchenko A. R., Shoemaker B. A., Thiessen P. A., Geer L. Y., Bryant S. H. ( 2002). CDD: a database of conserved domain alignments with links to domain three-dimensional structure. Nucleic Acids Res 30:281–283 [View Article][PubMed]
    [Google Scholar]
  27. Marrakchi H., Zhang Y. M., Rock C. O. ( 2002). Mechanistic diversity and regulation of type II fatty acid synthesis. Biochem Soc Trans 30:1050–1055 [View Article][PubMed]
    [Google Scholar]
  28. Matsuoka H., Hirooka K., Fujita Y. ( 2007). Organization and function of the YsiA regulon of Bacillus subtilis involved in fatty acid degradation. J Biol Chem 282:5180–5194 [View Article][PubMed]
    [Google Scholar]
  29. Orth P., Schnappinger D., Hillen W., Saenger W., Hinrichs W. ( 2000). Structural basis of gene regulation by the tetracycline inducible Tet repressor-operator system. Nat Struct Biol 7:215–219 [View Article][PubMed]
    [Google Scholar]
  30. Oshima T., Imahori K. ( 1974). Description of Thermus thermophilus (Yoshida and Oshima) com. nov., a non-sporulating thermophilic bacterium from a Japanese thermal spa. Int J Syst Bacteriol 24:102–112 [View Article]
    [Google Scholar]
  31. Oshima M., Miyagawa A. ( 1974). Comparative studies on the fatty acid composition of moderately and extremely thermophilic bacteria. Lipids 9:476–480 [View Article][PubMed]
    [Google Scholar]
  32. Otwinowski Z., Minor W. ( 1997). Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol 276:307–326 [View Article]
    [Google Scholar]
  33. Pauli G., Ehring R., Overath P. ( 1974). Fatty acid degradation in Escherichia coli: requirement of cyclic adenosine monophosphate and cyclic adenosine monophosphate receptor protein for enzyme synthesis. J Bacteriol 117:1178–1183[PubMed]
    [Google Scholar]
  34. Pepper S. D., Saunders E. K., Edwards L. E., Wilson C. L., Miller C. J. ( 2007). The utility of MAS5 expression summary and detection call algorithms. BMC Bioinformatics 8:273 [View Article][PubMed]
    [Google Scholar]
  35. Perrakis A., Harkiolaki M., Wilson K. S., Lamzin V. S. ( 2001). ARP/wARP and molecular replacement. Acta Crystallogr D Biol Crystallogr 57:1445–1450 [View Article][PubMed]
    [Google Scholar]
  36. Rock C. O., Cronan J. E. ( 1996). Escherichia coli as a model for the regulation of dissociable (type II) fatty acid biosynthesis. Biochim Biophys Acta 1302:1–16[PubMed] [CrossRef]
    [Google Scholar]
  37. Sakamoto K., Agari Y., Agari K., Kuramitsu S., Shinkai A. ( 2010). Structural and functional characterization of the transcriptional repressor CsoR from Thermus thermophilus HB8. Microbiology 156:1993–2005 [View Article][PubMed]
    [Google Scholar]
  38. Sanger F., Nicklen S., Coulson A. R. ( 1977). DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A 74:5463–5467 [View Article][PubMed]
    [Google Scholar]
  39. Schneider T. D., Stephens R. M. ( 1990). Sequence logos: a new way to display consensus sequences. Nucleic Acids Res 18:6097–6100 [View Article][PubMed]
    [Google Scholar]
  40. Schujman G. E., Paoletti L., Grossman A. D., de Mendoza D. ( 2003). FapR, a bacterial transcription factor involved in global regulation of membrane lipid biosynthesis. Dev Cell 4:663–672 [View Article][PubMed]
    [Google Scholar]
  41. Schujman G. E., Guerin M., Buschiazzo A., Schaeffer F., Llarrull L. I., Reh G., Vila A. J., Alzari P. M., de Mendoza D. ( 2006). Structural basis of lipid biosynthesis regulation in Gram-positive bacteria. EMBO J 25:4074–4083 [View Article][PubMed]
    [Google Scholar]
  42. Schumacher M. A., Miller M. C., Grkovic S., Brown M. H., Skurray R. A., Brennan R. G. ( 2001). Structural mechanisms of QacR induction and multidrug recognition. Science 294:2158–2163 [View Article][PubMed]
    [Google Scholar]
  43. Shinkai A., Kira S., Nakagawa N., Kashihara A., Kuramitsu S., Yokoyama S. ( 2007). Transcription activation mediated by a cyclic AMP receptor protein from Thermus thermophilus HB8. J Bacteriol 189:3891–3901 [View Article][PubMed]
    [Google Scholar]
  44. Siristova L., Melzoch K., Rezanka T. ( 2009). Fatty acids, unusual glycophospholipids and DNA analyses of thermophilic bacteria isolated from hot springs. Extremophiles 13:101–109 [View Article][PubMed]
    [Google Scholar]
  45. Storey J. D. ( 2002). A direct approach to false discovery rates. J R Stat Soc, B 64:479–498 [View Article]
    [Google Scholar]
  46. Storey J. D., Tibshirani R. ( 2003). Statistical significance for genomewide studies. Proc Natl Acad Sci U S A 100:9440–9445 [View Article][PubMed]
    [Google Scholar]
  47. Subrahmanyam S., Cronan J. E. Jr ( 1998). Overproduction of a functional fatty acid biosynthetic enzyme blocks fatty acid synthesis in Escherichia coli . J Bacteriol 180:4596–4602[PubMed]
    [Google Scholar]
  48. Terwilliger T. C., Berendzen J. ( 1999). Automated MAD and MIR structure solution. Acta Crystallogr D Biol Crystallogr 55:849–861 [View Article][PubMed]
    [Google Scholar]
  49. Ueno G., Kanda H., Hirose R., Ida K., Kumasaka T., Yamamoto M. ( 2006). RIKEN structural genomics beamlines at the SPring-8; high throughput protein crystallography with automated beamline operation. J Struct Funct Genomics 7:15–22 [View Article][PubMed]
    [Google Scholar]
  50. van Aalten D. M., DiRusso C. C., Knudsen J. ( 2001). The structural basis of acyl coenzyme A-dependent regulation of the transcription factor FadR. EMBO J 20:2041–2050 [View Article][PubMed]
    [Google Scholar]
  51. Willems A. R., Tahlan K., Taguchi T., Zhang K., Lee Z. Z., Ichinose K., Junop M. S., Nodwell J. R. ( 2008). Crystal structures of the Streptomyces coelicolor TetR-like protein ActR alone and in complex with actinorhodin or the actinorhodin biosynthetic precursor (S)-DNPA. J Mol Biol 376:1377–1387 [View Article][PubMed]
    [Google Scholar]
  52. Zhang Y. M., Marrakchi H., Rock C. O. ( 2002). The FabR (YijC) transcription factor regulates unsaturated fatty acid biosynthesis in Escherichia coli . J Biol Chem 277:15558–15565 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.048017-0
Loading
/content/journal/micro/10.1099/mic.0.048017-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Supplementary material 2

PDF

Supplementary material 3

PDF

Supplementary material 4

PDF

Supplementary material 5

PDF
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error