Advertisement

Current Microbiology

, Volume 66, Issue 4, pp 331–336 | Cite as

Investigating the Role of Protein UnkG from the Pseudomonas putida UW4 in the Ability of the Bacterium to Facilitate Plant Growth

  • Wei Jiang
  • Zhenyu Cheng
  • Brendan J. McConkey
  • Bernard R. Glick
Article

Abstract

A previous study showed that overexpressing protein UnkG decreased the ability of the plant growth-promoting bacterium Pseudomonas putida UW4 to facilitate plant growth and an unkG knockout mutant of P. putida UW4 displayed increased plant growth promotion. When activities of wild-type and the UnkG overexpressing strain, including growth rates, carbon utilization, cell size, 3-indoleacetic acid production, and 1-aminocyclopropane-1-carboxylate deaminase activity, were measured, there were no apparent differences between the strains. Monitoring proteome-level changes to the wild-type and overexpressing transformant by means of two-dimensional difference in-gel electrophoresis followed by mass spectrometry identification of the altered proteins, 1839 protein spots were detected and 16 of the 84 protein spots with changed expression levels were identified. Proteins with increased expression included arginine deiminase, dihydrodipicolinate synthase, azurin, flavoprotein (α-subunit), ferredoxin-NADP reductase, ATP-dependent Hs1 protease (ATP-binding subunit), UDP-N-acetyl muramate-l-alanine ligase, biotin carboxyl carrier protein subunit of acetyl-CoA carboxylase, and Fis two-component transcriptional regulator. Proteins with decreased expression included glutaminase-asparaginase, arginine/ornithine ABC transporter, cell division protein FtsZ and glutamyl-tRNA synthetase. The functions of three of the 16 proteins could not be identified. The results are consistent with UnkG being detrimental to plant growth because it acts as a regulatory protein that negatively affects several key cellular functions related to the energy balance of the bacterium.

Keywords

Protein Spot Pseudomonas Putida Flavin Adenine Dinucleotide Flavin Adenine Dinucleotide Arginine Deiminase 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

This work was supported by the Natural Science and Engineering Research Council of Canada Grant to B.R.G. We thank Mr. Dale Weber, Department of Biology, University of Waterloo, for help with electron microscopy.

Supplementary material

284_2012_279_MOESM1_ESM.docx (54 kb)
Supplementary material 1 (DOCX 54 kb)
284_2012_279_MOESM2_ESM.docx (556 kb)
Supplementary material 2 (DOCX 556 kb)
284_2012_279_MOESM3_ESM.docx (114 kb)
Supplementary material 3 (DOCX 114 kb)
284_2012_279_MOESM4_ESM.docx (71 kb)
Supplementary material 4 (DOCX 71 kb)

References

  1. 1.
    Aliverti A, Pandini V, Pennati A, de Rosa M, Zanetti G (2008) Structural and functional diversity of ferredoxin-NADP(+) reductases. Arch Biochem Biophys 474:283–291PubMedCrossRefGoogle Scholar
  2. 2.
    Bochtler M, Hartmann C, Song HK, Bourenkov GP, Bartunik HD, Huber R (2000) The structures of HslU and the ATP-dependent protease HslU-HslV. Science 403:800–805Google Scholar
  3. 3.
    Brownsey RW, Boone AN, Elliott JE, Kulpa JE, Lee WM (2006) Regulation of acetyl-CoA carboxylase. Biochem Soc Trans 34:223–227PubMedCrossRefGoogle Scholar
  4. 4.
    Chen Z, Barber MJ, McIntire WS, Mathews FS (1998) Crystallographic study of azurin from Pseudomonas putida. Acta Cryst D54:253–268Google Scholar
  5. 5.
    Cheng Z, Duan J, Hao Y, McConkey BJ, Glick BR (2009) Identification of bacterial proteins mediating the interactions between P. putida UW4 and Brassica napus (canola). Mol Plant-Microbe Interact 22:686–694PubMedCrossRefGoogle Scholar
  6. 6.
    Cheng Z, Woody OZ, Song J, Glick BR, McConkey BJ (2009) Proteome reference map for the plant growth-promoting bacterium Pseudomonas putida UW4. Proteomics 9:4271–4274PubMedCrossRefGoogle Scholar
  7. 7.
    Edgar RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 32:1792–1797PubMedCrossRefGoogle Scholar
  8. 8.
    Emanuele JJ Jr, Jin H, Jacobson BL, Chang CY, Einspahr HM, Villafranca JJ (1996) Kinetic and crystallographic studies of Escherichia coli UDP-N-acetylmuramate: l-alanine ligase. Protein Sci 5:2566–2574PubMedCrossRefGoogle Scholar
  9. 9.
    Fernández-Piñar R, Ramos JL, Rodríguez-Herva JJ, Espinosa-Urgel M (2008) A two-component regulatory system integrates redox state and population density sensing in Pseudomonas putida. J Bacteriol 190:7666–7674PubMedCrossRefGoogle Scholar
  10. 10.
    Glick BR (2005) Modulation of plant ethylene levels by the enzyme ACC deaminase. FEMS Microbiol Lett 251:1–7PubMedCrossRefGoogle Scholar
  11. 11.
    Glick BR, Cheng Z, Czarny J, Duan J (2007) Promotion of plant growth by ACC deaminase-containing soil bacteria. Eur J Plant Pathol 119:329–339CrossRefGoogle Scholar
  12. 12.
    Glick BR, Todorovic B, Czarny J, Cheng Z, Duan J, McConkey B (2007) Promotion of plant growth by bacterial ACC deaminase. Crit Rev Plant Sci 26:227–242CrossRefGoogle Scholar
  13. 13.
    Guindon S, Gascuel O (2003) A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol 52:696–704PubMedCrossRefGoogle Scholar
  14. 14.
    Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for windows 95/98/NT. Nucleic Acids Symp Ser 41:95–98Google Scholar
  15. 15.
    Jakovleva J, Teppo A, Velts A, Saumaa S, Moor H, Kivisaar M, Teras R (2012) Fis regulates the competitiveness of Pseudomonas putida on barley roots by inducing biofilm formation. Microbiology 158:708–720PubMedCrossRefGoogle Scholar
  16. 16.
    Kaur N, Gautam A, Kumar S, Singh A, Singh N, Sharma S, Sharma R, Tewari R, Singh TP (2011) Biochemical studies and crystal structure determination of dihydrodipicolinate synthase from Pseudononas aeruginosa. Int J Biol Macromol 48:779–787PubMedCrossRefGoogle Scholar
  17. 17.
    Lu X, Galkin A, Herzberg O, Dunaway-Mariano D (2004) Arginine deiminase uses an active-site cysteine in nucleophilic catalysis of l-arginine hydrolysis. J Am Chem Soc 126:5374–5375PubMedCrossRefGoogle Scholar
  18. 18.
    Ma B, Zhang K, Hendrie C, Liang C, Li M, Doherty-Kirby A, Lajoie G (2003) PEAKS: powerful software for peptide de novo sequencing by tandem mass spectrometry. Rapid Commun Mass Spectrom 17:2337–2342PubMedCrossRefGoogle Scholar
  19. 19.
    Oliva MA, Trambaiolo D, Löwe J (2007) Structural insights into the conformational variability of FtsZ. J Mol Biol 373:1229–1242PubMedCrossRefGoogle Scholar
  20. 20.
    Ortlund E, Lacount MW, Lewinski K, Lebioda L (2000) Reactions of Pseudomonas 7A glutaminase-asparaginase with diazo analogues of glutamine and asparagines result in unexpected covalent inhibitions and suggests an unusual catalytic triad Thr-Tyr-Glu. Biochemistry 39:1199–1204PubMedCrossRefGoogle Scholar
  21. 21.
    Patten CL, Glick BR (2002) Role of Pseudomonas putida indoleacetic acid in development of the host plant root system. Appl Environ Microbiol 68:3795–3801PubMedCrossRefGoogle Scholar
  22. 22.
    Penrose DM, Glick BR (2003) Methods for isolating and characterizing ACC deaminase-containing plant growth-promoting rhizobacteria. Physiol Plant 118:10–15PubMedCrossRefGoogle Scholar
  23. 23.
    Perkins DN, Pappin DJ, Creasy DM, Cottrell JS (1999) Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20:3551–3567PubMedCrossRefGoogle Scholar
  24. 24.
    Roberts DL, Salazar D, Fulmer JP, Frerman FE, Kim JJP (1999) Crystal structure of Paracoccus denitrificans electron transfer flavoprotein and electrostatic analysis of a conserved flavin binding domain. Biochemistry 38:1977–1989PubMedCrossRefGoogle Scholar
  25. 25.
    Rohrwild M, Coux O, Huang HC, Moerschell RP, Yoo SJ, Seol JH, Chung CH, Goldberg AL (1996) HslV-HslU: a novel ATP-dependent protease complex in Escherichia coli related to the eukaryotic proteasome. Proc Natl Acad Sci USA 93:5808–5813PubMedCrossRefGoogle Scholar
  26. 26.
    Yamao F, Inokuchi H, Cheung A, Ozeki H, Soell DG (1982) Escherichia coli glutaminyl-tRNA synthetase I. Isolation and DNA sequence of the glnS gene. J Biol Chem 257:11639–11643PubMedGoogle Scholar
  27. 27.
    Yeom J, Jeon CO, Madsen EL, Park W (2009) Ferredoxin-NADP+ reductase from Pseudomonas putida functions asa ferric reductase. J Bacteriol 191:1472–1479PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2012

Authors and Affiliations

  • Wei Jiang
    • 1
  • Zhenyu Cheng
    • 1
  • Brendan J. McConkey
    • 1
  • Bernard R. Glick
    • 1
  1. 1.Department of BiologyUniversity of WaterlooWaterlooCanada

Personalised recommendations