1887

Abstract

CbbR is a LysR-type transcriptional regulator that activates expression of the operons containing () genes that encode the CO fixation pathway enzymes in () under autotrophic growth conditions. The operons are stringently downregulated during chemoheterotrophic growth on organic acids such as malate. CbbR constitutive proteins (CbbR*s), typically with single amino acid substitutions, were selected and isolated that activate expression of the operons under chemoheterotrophic growth conditions. A large set of CbbR*s from all major domains of the CbbR molecule were identified, except for the DNA-binding domain. The level of gene expression conferred for many of these CbbR*s under autotrophic growth was greater than that conferred by wild-type CbbR. Several of these CbbR*s increase transcription two- to threefold more than wild-type CbbR. One particular CbbR*, a truncated protein, was useful in identifying the regions of CbbR that are necessary for transcriptional activation and, by logical extension, necessary for interaction with RNA polymerase. The reductive assimilation of carbon via CO fixation is an important step in the cost-effective production of useful biological compounds. Enhancing CO fixation in through greater transcriptional activation of the operons could prove advantageous, and the use of CbbR*s is one way to enhance product formation.

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2015-09-01
2019-12-08
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References

  1. Akakura R., Winans S. C.. ( 2002;). Constitutive mutations of the OccR regulatory protein affect DNA bending in response to metabolites released from plant tumors. J Biol Chem 277: 5866–5874 [CrossRef] [PubMed].
    [Google Scholar]
  2. Balcewich M. D., Reeve T. M., Orlikow E. A., Donald L. J., Vocadlo D. J., Mark B. L.. ( 2010;). Crystal structure of the AmpR effector binding domain provides insight into the molecular regulation of inducible AmpC β-lactamase. J Mol Biol 400: 998–1010 [CrossRef] [PubMed].
    [Google Scholar]
  3. Bartowsky E., Normark S.. ( 1991;). Purification and mutant analysis of Citrobacter freundii AmpR, the regulator for chromosomal AmpC β-lactamase. Mol Microbiol 5: 1715–1725 [CrossRef] [PubMed].
    [Google Scholar]
  4. Batchelor J. D., Lee P. S., Wang A. C., Doucleff M., Wemmer D. E.. ( 2013;). Structural mechanism of GAF-regulated σ(54) activators from Aquifex aeolicus. J Mol Biol 425: 156–170 [CrossRef] [PubMed].
    [Google Scholar]
  5. Belitsky B. R., Sonenshein A. L.. ( 1997;). Altered transcription activation specificity of a mutant form of Bacillus subtilis GltR, a LysR family member. J Bacteriol 179: 1035–1043 [PubMed].
    [Google Scholar]
  6. Bender R. A.. ( 1991;). The role of the NAC protein in the nitrogen regulation of Klebsiella aerogenes. Mol Microbiol 5: 2575–2580 [CrossRef] [PubMed].
    [Google Scholar]
  7. Bowien B., Kusian B.. ( 2002;). Genetics and control of CO2 assimilation in the chemoautotroph Ralstonia eutropha. Arch Microbiol 178: 85–93 [CrossRef] [PubMed].
    [Google Scholar]
  8. Bowien B., Friedrich B., Friedrich C. G.. ( 1984;). Involvement of megaplasmids in heterotrophic derepression of the carbon dioxide-assimilating enzyme system in Alcaligenes spp. Arch Microbiol 139: 305–310 [CrossRef].
    [Google Scholar]
  9. Bowien B., Windhövel U., Yoo J.-G., Bednarski R., Kusian B.. ( 1990;). Genetics of CO2 fixation in the chemoautotroph Alcaligenes eutrophus. FEMS Microbiol Rev 87: 445–450 [CrossRef].
    [Google Scholar]
  10. Burn J. E., Hamilton W. D., Wootton J. C., Johnston A. W.. ( 1989;). Single and multiple mutations affecting properties of the regulatory gene nodD of Rhizobium. Mol Microbiol 3: 1567–1577 [CrossRef] [PubMed].
    [Google Scholar]
  11. Bykowski T., van der Ploeg J. R., Iwanicka-Nowicka R., Hryniewicz M. M.. ( 2002;). The switch from inorganic to organic sulphur assimilation in Escherichia coli: adenosine 5′-phosphosulphate (APS) as a signalling molecule for sulphate excess. Mol Microbiol 43: 1347–1358 [CrossRef] [PubMed].
    [Google Scholar]
  12. Cebolla A., Sousa C., de Lorenzo V.. ( 1997;). Effector specificity mutants of the transcriptional activator NahR of naphthalene degrading Pseudomonas define protein sites involved in binding of aromatic inducers. J Biol Chem 272: 3986–3992 [CrossRef] [PubMed].
    [Google Scholar]
  13. Choi H., Kim S., Mukhopadhyay P., Cho S., Woo J., Storz G., Ryu S. E.. ( 2001;). Structural basis of the redox switch in the OxyR transcription factor. Cell 105: 103–113 [CrossRef] [PubMed].
    [Google Scholar]
  14. Colyer T. E., Kredich N. M.. ( 1994;). Residue threonine-149 of the Salmonella typhimurium CysB transcription activator: mutations causing constitutive expression of positively regulated genes of the cysteine regulon. Mol Microbiol 13: 797–805 [CrossRef] [PubMed].
    [Google Scholar]
  15. Dangel A. W., Tabita F. R.. ( 2009;). Protein-protein interactions between CbbR and RegA (PrrA), transcriptional regulators of the cbb operons of Rhodobacter sphaeroides. Mol Microbiol 71: 717–729 [CrossRef] [PubMed].
    [Google Scholar]
  16. Dangel A. W., Gibson J. L., Janssen A. P., Tabita F. R.. ( 2005;). Residues that influence in vivo and in vitro CbbR function in Rhodobacter sphaeroides and identification of a specific region critical for co-inducer recognition. Mol Microbiol 57: 1397–1414 [CrossRef] [PubMed].
    [Google Scholar]
  17. Dangel A. W., Luther A., Tabita F. R.. ( 2014;). Amino acid residues of RegA important for interactions with the CbbR-DNA complex of Rhodobacter sphaeroides. J Bacteriol 196: 3179–3190 [CrossRef] [PubMed].
    [Google Scholar]
  18. Dubbs J. M., Tabita F. R.. ( 1998;). Two functionally distinct regions upstream of the cbb I operon of Rhodobacter sphaeroides regulate gene expression. J Bacteriol 180: 4903–4911 [PubMed].
    [Google Scholar]
  19. Dubbs P., Dubbs J. M., Tabita F. R.. ( 2004;). Effector-mediated interaction of CbbRI and CbbRII regulators with target sequences in Rhodobacter capsulatus. J Bacteriol 186: 8026–8035 [CrossRef] [PubMed].
    [Google Scholar]
  20. Ezezika O. C., Haddad S., Clark T. J., Neidle E. L., Momany C.. ( 2007;). Distinct effector-binding sites enable synergistic transcriptional activation by BenM, a LysR-type regulator. J Mol Biol 367: 616–629 [CrossRef] [PubMed].
    [Google Scholar]
  21. Falcone D. L., Quivey R. G. Jr, Tabita F. R.. ( 1988;). Transposon mutagenesis and physiological analysis of strains containing inactivated form I and form II ribulose bisphosphate carboxylase/oxygenase genes in Rhodobacter sphaeroides. J Bacteriol 170: 5–11 [PubMed].
    [Google Scholar]
  22. Figurski D. H., Helinski D. R.. ( 1979;). Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc Natl Acad Sci U S A 76: 1648–1652 [CrossRef] [PubMed].
    [Google Scholar]
  23. Friedrich C. G., Bowien B., Friedrich B.. ( 1979;). Formate and oxalate metabolism in Alcaligenes eutrophus. J Gen Microbiol 115: 185–192 [CrossRef].
    [Google Scholar]
  24. Gibson J. L., Tabita F. R.. ( 1977;). Different molecular forms of d-ribulose-1,5-bisphosphate carboxylase from Rhodopseudomonas sphaeroides. J Biol Chem 252: 943–949 [PubMed].
    [Google Scholar]
  25. Gibson J. L., Tabita F. R.. ( 1993;). Nucleotide sequence and functional analysis of cbbR, a positive regulator of the Calvin cycle operons of Rhodobacter sphaeroides. J Bacteriol 175: 5778–5784 [PubMed].
    [Google Scholar]
  26. Grzeszik C., Jeffke T., Schäferjohann J., Kusian B., Bowien B.. ( 2000;). Phosphoenolpyruvate is a signal metabolite in transcriptional control of the cbb CO2 fixation operons in Ralstonia eutropha. J Mol Microbiol Biotechnol 2: 311–320 [PubMed].
    [Google Scholar]
  27. Hayes R. P., Moural T. W., Lewis K. M., Onofrei D., Xun L., Kang C.. ( 2014;). Structures of the inducer-binding domain of pentachlorophenol-degrading gene regulator PcpR from Sphingobium chlorophenolicum. Int J Mol Sci 15: 20736–20752 [CrossRef] [PubMed].
    [Google Scholar]
  28. Hou B., Li F., Yang X., Hong G.. ( 2009;). A small functional intramolecular region of NodD was identified by mutation. Acta Biochim Biophys Sin (Shanghai) 41: 822–830 [CrossRef] [PubMed].
    [Google Scholar]
  29. Jørgensen C., Dandanell G.. ( 1999;). Isolation and characterization of mutations in the Escherichia coli regulatory protein XapR. J Bacteriol 181: 4397–4403 [PubMed].
    [Google Scholar]
  30. Joshi G. S., Zianni M., Bobst C. E., Tabita F. R.. ( 2012;). Further unraveling the regulatory twist by elucidating metabolic coinducer-mediated CbbR-cbbI promoter interactions in Rhodopseudomonas palustris CGA010. J Bacteriol 194: 1350–1360 [CrossRef] [PubMed].
    [Google Scholar]
  31. Joshi G. S., Zianni M., Bobst C. E., Tabita F. R.. ( 2013;). Regulatory twist and synergistic role of metabolic coinducer- and response regulator-mediated CbbR-cbb I interactions in Rhodopseudomonas palustris CGA010. J Bacteriol 195: 1381–1388 [CrossRef] [PubMed].
    [Google Scholar]
  32. Knauf V. C., Nester E. W.. ( 1982;). Wide host range cloning vectors: a cosmid clone bank of an Agrobacterium Ti plasmid. Plasmid 8: 45–54 [CrossRef] [PubMed].
    [Google Scholar]
  33. Kullik I., Toledano M. B., Tartaglia L. A., Storz G.. ( 1995;). Mutational analysis of the redox-sensitive transcriptional regulator OxyR: regions important for oxidation and transcriptional activation. J Bacteriol 177: 1275–1284 [PubMed].
    [Google Scholar]
  34. Kusian B., Bowien B.. ( 1995;). Operator binding of the CbbR protein, which activates the duplicate cbb CO2 assimilation operons of Alcaligenes eutrophus. J Bacteriol 177: 6568–6574 [PubMed].
    [Google Scholar]
  35. Kusian B., Bednarski R., Husemann M., Bowien B.. ( 1995;). Characterization of the duplicate ribulose-1,5-bisphosphate carboxylase genes and cbb promoters of Alcaligenes eutrophus. J Bacteriol 177: 4442–4450 [PubMed].
    [Google Scholar]
  36. Laguri C., Stenzel R. A., Donohue T. J., Phillips-Jones M. K., Williamson M. P.. ( 2006;). Activation of the global gene regulator PrrA (RegA) from Rhodobacter sphaeroides. Biochemistry 45: 7872–7881 [CrossRef] [PubMed].
    [Google Scholar]
  37. Lee J. H., Park D. O., Park S. W., Hwang E. H., Oh J. I., Kim Y. M.. ( 2009;). Expression and regulation of ribulose 1,5-bisphosphate carboxylase/oxygenase genes in Mycobacterium sp. strain JC1 DSM 3803. J Microbiol 47: 297–307 [CrossRef] [PubMed].
    [Google Scholar]
  38. Lochowska A., Iwanicka-Nowicka R., Plochocka D., Hryniewicz M. M.. ( 2001;). Functional dissection of the LysR-type CysB transcriptional regulator. Regions important for DNA binding, inducer response, oligomerization, and positive control. J Biol Chem 276: 2098–2107 [CrossRef] [PubMed].
    [Google Scholar]
  39. Maddocks S. E., Oyston P. C. F.. ( 2008;). Structure and function of the LysR-type transcriptional regulator (LTTR) family proteins. Microbiology 154: 3609–3623 [CrossRef] [PubMed].
    [Google Scholar]
  40. Miller J. H.. ( 1992;). A Short Course in Bacterial Genetics Cold Spring Harbor. NY: Cold Spring Harbor Laboratory Press;.
    [Google Scholar]
  41. Minoda A., Weber A. P. M., Tanaka K., Miyagishima S. Y.. ( 2010;). Nucleus-independent control of the rubisco operon by the plastid-encoded transcription factor Ycf30 in the red alga Cyanidioschyzon merolae. Plant Physiol 154: 1532–1540 [CrossRef] [PubMed].
    [Google Scholar]
  42. Monferrer D., Tralau T., Kertesz M. A., Dix I., Solà M., Usón I.. ( 2010;). Structural studies on the full-length LysR-type regulator TsaR from Comamonas testosteroni T-2 reveal a novel open conformation of the tetrameric LTTR fold. Mol Microbiol 75: 1199–1214 [CrossRef] [PubMed].
    [Google Scholar]
  43. Muraoka S., Okumura R., Ogawa N., Nonaka T., Miyashita K., Senda T.. ( 2003;). Crystal structure of a full-length LysR-type transcriptional regulator, CbnR: unusual combination of two subunit forms and molecular bases for causing and changing DNA bend. J Mol Biol 328: 555–566 [CrossRef] [PubMed].
    [Google Scholar]
  44. Muse W. B., Bender R. A.. ( 1999;). The amino-terminal 100 residues of the nitrogen assimilation control protein (NAC) encode all known properties of NAC from Klebsiella aerogenes and Escherichia coli. J Bacteriol 181: 934–940 [PubMed].
    [Google Scholar]
  45. Ormerod J. G., Ormerod K. S., Gest H.. ( 1961;). Light-dependent utilization of organic compounds and photoproduction of molecular hydrogen by photosynthetic bacteria; relationships with nitrogen metabolism. Arch Biochem Biophys 94: 449–463 [CrossRef] [PubMed].
    [Google Scholar]
  46. Romagnoli S., Tabita F. R.. ( 2006;). A novel three-protein two-component system provides a regulatory twist on an established circuit to modulate expression of the cbbI region of Rhodopseudomonas palustris CGA010. J Bacteriol 188: 2780–2791 [CrossRef] [PubMed].
    [Google Scholar]
  47. Ruangprasert A., Craven S. H., Neidle E. L., Momany C.. ( 2010;). Full-length structures of BenM and two variants reveal different oligomerization schemes for LysR-type transcriptional regulators. J Mol Biol 404: 568–586 [CrossRef] [PubMed].
    [Google Scholar]
  48. Sainsbury S., Lane L. A., Ren J., Gilbert R. J., Saunders N. J., Robinson C. V., Stuart D. I., Owens R. J.. ( 2009;). The structure of CrgA from Neisseria meningitidis reveals a new octameric assembly state for LysR transcriptional regulators. Nucleic Acids Res 37: 4545–4558 [CrossRef] [PubMed].
    [Google Scholar]
  49. Schell M. A.. ( 1993;). Molecular biology of the LysR family of transcriptional regulators. Annu Rev Microbiol 47: 597–626 [CrossRef] [PubMed].
    [Google Scholar]
  50. Schwacha A., Bender R. A.. ( 1993;). The product of the Klebsiella aerogenes nac (nitrogen assimilation control) gene is sufficient for activation of the hut operons and repression of the gdh operon. J Bacteriol 175: 2116–2124 [PubMed].
    [Google Scholar]
  51. Shapira S. K., Chou J., Richaud F. V., Casadaban M. J.. ( 1983;). New versatile plasmid vectors for expression of hybrid proteins coded by a cloned gene fused to lacZ gene sequences encoding an enzymatically active carboxy-terminal portion of β-galactosidase. Gene 25: 71–82 [CrossRef] [PubMed].
    [Google Scholar]
  52. Smirnova I. A., Dian C., Leonard G. A., McSweeney S., Birse D., Brzezinski P.. ( 2004;). Development of a bacterial biosensor for nitrotoluenes: the crystal structure of the transcriptional regulator DntR. J Mol Biol 340: 405–418 [CrossRef] [PubMed].
    [Google Scholar]
  53. Smith S. A., Tabita F. R.. ( 2002;). Up-regulated expression of the cbb I cbb II operons during photoheterotrophic growth of a ribulose 1,5-bisphosphate carboxylase-oxygenase deletion mutant of Rhodobacter sphaeroides. J Bacteriol 184: 6721–6724 [CrossRef] [PubMed].
    [Google Scholar]
  54. Stec E., Witkowska-Zimny M., Hryniewicz M. M., Neumann P., Wilkinson A. J., Brzozowski A. M., Verma C. S., Zaim J., Wysocki S., Bujacz G. D.. ( 2006;). Structural basis of the sulphate starvation response in E. coli: crystal structure and mutational analysis of the cofactor-binding domain of the Cbl transcriptional regulator. J Mol Biol 364: 309–322 [CrossRef] [PubMed].
    [Google Scholar]
  55. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W.. ( 1990;). Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol 185: 60–89 [CrossRef] [PubMed].
    [Google Scholar]
  56. Taylor J. L., De Silva R. S., Kovacikova G., Lin W., Taylor R. K., Skorupski K., Kull F. J.. ( 2012;). The crystal structure of AphB, a virulence gene activator from Vibrio cholerae, reveals residues that influence its response to oxygen and pH. Mol Microbiol 83: 457–470 [CrossRef] [PubMed].
    [Google Scholar]
  57. Terazono K., Hayashi N. R., Igarashi Y.. ( 2001;). CbbR, a LysR-type transcriptional regulator from Hydrogenophilus thermoluteolus, binds two cbb promoter regions. FEMS Microbiol Lett 198: 151–157 [CrossRef] [PubMed].
    [Google Scholar]
  58. Tyrrell R., Verschueren K. H., Dodson E. J., Murshudov G. N., Addy C., Wilkinson A. J.. ( 1997;). The structure of the cofactor-binding fragment of the LysR family member, CysB: a familiar fold with a surprising subunit arrangement. Structure 5: 1017–1032 [CrossRef] [PubMed].
    [Google Scholar]
  59. Vadlamani G., Thomas M. D., Patel T. R., Donald L. J., Reeve T. M., Stetefeld J., Standing K. G., Vocadlo D. J., Mark B. L.. ( 2015;). The β-lactamase gene regulator AmpR is a tetramer that recognizes and binds the d-Ala-d-Ala motif of its repressor UDP-N-acetylmuramic acid (MurNAc)-pentapeptide. J Biol Chem 290: 2630–2643 [CrossRef] [PubMed].
    [Google Scholar]
  60. van Keulen G., Girbal L., van den Bergh E. R. E., Dijkhuizen L., Meijer W. G.. ( 1998;). The LysR-type transcriptional regulator CbbR controlling autotrophic CO2 fixation by Xanthobacter flavus is an NADPH sensor. J Bacteriol 180: 1411–1417 [PubMed].
    [Google Scholar]
  61. van Keulen G., Ridder A. N. J. A., Dijkhuizen L., Meijer W. G.. ( 2003;). Analysis of DNA binding and transcriptional activation by the LysR-type transcriptional regulator CbbR of Xanthobacter flavus. J Bacteriol 185: 1245–1252 [CrossRef] [PubMed].
    [Google Scholar]
  62. Viale A. M., Kobayashi H., Akazawa T., Henikoff S.. ( 1991;). rbcR, a gene coding for a member of the LysR family of transcriptional regulators, is located upstream of the expressed set of ribulose 1,5-bisphosphate carboxylase/oxygenase genes in the photosynthetic bacterium Chromatium vinosum. J Bacteriol 173: 5224–5229 [PubMed].
    [Google Scholar]
  63. Voss I., Steinbüchel A.. ( 2006;). Application of a KDPG-aldolase gene-dependent addiction system for enhanced production of cyanophycin in Ralstonia eutropha strain H16. Metab Eng 8: 66–78 [CrossRef] [PubMed].
    [Google Scholar]
  64. Windhövel U., Bowien B.. ( 1990;). On the operon structure of the cfx gene clusters in Alcaligenes eutrophus. . Arch Microbiol 154, 85–91. [PubMed].
  65. Windhövel U., Bowien B.. ( 1991;). Identification of cfxR, an activator gene of autotrophic CO2 fixation in Alcaligenes eutrophus. Mol Microbiol 5: 2695–2705 [CrossRef] [PubMed].
    [Google Scholar]
  66. Yanisch-Perron C., Vieira J., Messing J.. ( 1985;). Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene 33: 103–119 [CrossRef] [PubMed].
    [Google Scholar]
  67. Zhou X., Lou Z., Fu S., Yang A., Shen H., Li Z., Feng Y., Bartlam M., Wang H., Rao Z.. ( 2010;). Crystal structure of ArgP from Mycobacterium tuberculosis confirms two distinct conformations of full-length LysR transcriptional regulators and reveals its function in DNA binding and transcriptional regulation. J Mol Biol 396: 1012–1024 [CrossRef] [PubMed].
    [Google Scholar]
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