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Evidence for vital role of endo-β-N-acetylglucosaminidase in the resistance of Arthrobacter protophormiae RKJ100 towards elevated concentrations of o-nitrobenzoate

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Abstract

Arthrobacter protophormiae RKJ100 was previously characterized for its ability to tolerate extremely high concentrations of o-nitrobenzoate (ONB), a toxic xenobiotic environmental pollutant. The physiological responses of strain RKJ100 to ≥30 mM ONB indicated towards a resistance mechanism manifested via alteration of cell morphology and cell wall structure. In this study, we aim to characterize gene(s) involved in the resistance of strain RKJ100 towards extreme concentrations (i.e. 150 mM) of ONB. Transposon mutagenesis was carried out to generate a mutant library of strain RKJ100, which was then screened for ONB-sensitive mutants. A sensitive mutant was defined and selected as one that could not tolerate ≥30 mM ONB. Molecular and biochemical characterization of this mutant showed that the disruption of endo-β-N-acetylglucosaminidase (ENGase) gene caused the sensitivity. ENGase is an important enzyme for oligosaccharide processing and cell wall recycling in bacteria, fungi, plants and animals. Previous reports have already indicated several possible roles of this enzyme in cellular homeostasis. Results presented here provide the first evidence for its involvement in bacterial resistance towards extreme concentrations of a toxic xenobiotic compound and also suggest that strain RKJ100 employs ENGase as an important component in osmotic shock response for resisting extreme concentrations of ONB.

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References

  • Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Baldwin WW, Sheu MJ, Bankston PW, Woldringh CL (1988) Changes in buoyant density and cell size of Escherichia coli in response to osmotic shocks. J Bacteriol 170:452–455

    PubMed Central  PubMed  CAS  Google Scholar 

  • Bohin JP (2000) Osmoregulated periplasmic glucans in Proteobacteria. FEMS Microbiol Lett 186:11–19

    Article  PubMed  CAS  Google Scholar 

  • Bouchart F, Delangle A, Lemoine J, Bohin JP, Lacroix JM (2007) Proteomic analysis of a non-virulent mutant of the phytopathogenic bacterium Erwinia chrysanthemi deficient in osmoregulated periplasmic glucans: change in protein expression is not restricted to the envelope, but affects general metabolism. Microbiology 153:760–767

    Article  PubMed  CAS  Google Scholar 

  • Brown T (2001) Southern blotting. Curr Protoc Immunol, Chapter 10: Unit 10 16A

  • Chauhan A, Jain RK (2000) Degradation of o-nitrobenzoate via anthranilic acid (o-aminobenzoate) by Arthrobacter protophormiae: a plasmid-encoded new pathway. Biochem Biophys Res Commun 267:236–244

    Article  PubMed  CAS  Google Scholar 

  • Chauhan A, Chakraborti AK, Jain RK (2000) Plasmid-encoded degradation of p-nitrophenol and 4-nitrocatechol by Arthrobacter protophormiae. Biochem Biophys Res Commun 270:733–740

    Article  PubMed  CAS  Google Scholar 

  • Cheng Q, Li H, Merdek K, Park JT (2000) Molecular characterization of the beta-N-acetylglucosaminidase of Escherichia coli and its role in cell wall recycling. J Bacteriol 182:4836–4840

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Csonka LN (1989) Physiological and genetic responses of bacteria to osmotic stress. Microbiol Rev 53:121–147

    PubMed Central  PubMed  CAS  Google Scholar 

  • de Lorenzo V (2008) Systems biology approaches to bioremediation. Curr Opin Biotechnol 19:579–589

    Article  PubMed  CAS  Google Scholar 

  • Ensign JC (1970) Long-term starvation survival of rod and spherical cells of Arthrobacter crystallopoietes. J Bacteriol 103:569–577

    PubMed Central  PubMed  CAS  Google Scholar 

  • Fan JQ, Takegawa K, Iwahara S, Kondo A, Kato I, Abeygunawardana C, Lee YC (1995) Enhanced transglycosylation activity of Arthrobacter protophormiae endo-beta-N-acetylglucosaminidase in media containing organic solvents. J Biol Chem 270:17723–17729

    Article  PubMed  CAS  Google Scholar 

  • Fontana R, Canepari P, Satta G (1979) Alterations in peptidoglycan chemical composition associated with rod-to-sphere transition in a conditional mutant of Klebsiella pneumoniae. J Bacteriol 139:1028–1038

    PubMed Central  PubMed  CAS  Google Scholar 

  • Gartemann KH, Eichenlaub R (2001) Isolation and characterization of IS1409, an insertion element of 4-chlorobenzoate-degrading Arthrobacter sp. strain TM1, and development of a system for transposon mutagenesis. J Bacteriol 183:3729–3736

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Germida JJ, Casida LE Jr (1980) Myceloid growth of Arthrobacter globiformis and other Arthrobacter species. J Bacteriol 144:1152–1158

    PubMed Central  PubMed  CAS  Google Scholar 

  • Godoy P, Ramos-Gonzalez MI, Ramos JL (2001) Involvement of the TonB system in tolerance to solvents and drugs in Pseudomonas putida DOT-T1E. J Bacteriol 183:5285–5292

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Hauser S, Song H, Li H, Wang LX (2005) A novel fluorescence-based assay for the transglycosylation activity of endo-beta-N-acetylglucosaminidases. Biochem Biophys Res Commun 328:580–585

    Article  PubMed  CAS  Google Scholar 

  • Jordan S, Hutchings MI, Mascher T (2008) Cell envelope stress response in Gram-positive bacteria. FEMS Microbiol Rev 32:107–146

    Article  PubMed  CAS  Google Scholar 

  • Ling Z, Suits MD, Bingham RJ et al (2009) The X-ray crystal structure of an Arthrobacter protophormiae endo-beta-N-acetylglucosaminidase reveals a (beta/alpha)(8) catalytic domain, two ancillary domains and active site residues key for transglycosylation activity. J Mol Biol 389:1–9

    Article  PubMed  CAS  Google Scholar 

  • Liu YG, Whittier RF (1995) Thermal asymmetric interlaced PCR: automatable amplification and sequencing of insert end fragments from P1 and YAC clones for chromosome walking. Genomics 25:674–681

    Article  PubMed  CAS  Google Scholar 

  • Lovley DR (2003) Cleaning up with genomics: applying molecular biology to bioremediation. Nat Rev Microbiol 1:35–44

    Article  PubMed  CAS  Google Scholar 

  • Meganathan R, Ensign JC (1976) Stability of enzymes in starving Arthrobacter crystallopoietes. J Gen Microbiol 94:90–96

    Article  PubMed  CAS  Google Scholar 

  • Mongodin EF, Shapir N, Daugherty SC et al (2006) Secrets of soil survival revealed by the genome sequence of Arthrobacter aurescens TC1. PLoS Genet 2:e214

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Nakayama T, Soma M, Rahmutula D, Ozawa Y, Kanmatsuse K (2001) Isolation of the 5′-flanking region of genes by thermal asymmetric interlaced polymerase chain reaction. Med Sci Monit 7:345–349

    PubMed  CAS  Google Scholar 

  • Nothaft H, Liu X, McNally DJ, Li J, Szymanski CM (2009) Study of free oligosaccharides derived from the bacterial N-glycosylation pathway. Proc Natl Acad Sci USA 106:15019–15024

    Article  PubMed Central  PubMed  Google Scholar 

  • Pandey G, Paul D, Jain RK (2003) Branching of o-nitrobenzoate degradation pathway in Arthrobacter protophormiae RKJ100: identification of new intermediates. FEMS Microbiol Lett 229:231–236

    Article  PubMed  CAS  Google Scholar 

  • Pandey G, Pandey J, Jain RK (2006) Monitoring Arthrobacter protophormiae RKJ100 in a ‘tag and chase’ method during p-nitrophenol bio-remediation in soil microcosms. Appl Microbiol Biotechnol 70:757–760

    Article  PubMed  CAS  Google Scholar 

  • Pandey J, Chauhan A, Jain RK (2009) Integrative approaches for assessing the ecological sustainability of in situ bioremediation. FEMS Microbiol Rev 33:324–375

    Article  PubMed  CAS  Google Scholar 

  • Poolman B, Spitzer JJ, Wood JM (2004) Bacterial osmosensing: roles of membrane structure and electrostatics in lipid–protein and protein–protein interactions. Biochim Biophys Acta 1666:88–104

    Article  PubMed  CAS  Google Scholar 

  • Ramos JL, Duque E, Gallegos MT et al (2002) Mechanisms of solvent tolerance in gram-negative bacteria. Annu Rev Microbiol 56:743–768

    Article  PubMed  CAS  Google Scholar 

  • Ramos-Gonzalez MI, Godoy P, Alaminos M, Ben-Bassat A, Ramos JL (2001) Physiological characterization of Pseudomonas putida DOT-T1E tolerance to p-hydroxybenzoate. Appl Environ Microbiol 67:4338–4341

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Rodriguez-Herva JJ, Garcia V, Hurtado A, Segura A, Ramos JL (2007) The ttgGHI solvent efflux pump operon of Pseudomonas putida DOT-T1E is located on a large self-transmissible plasmid. Environ Microbiol 9:1550–1561

    Article  PubMed  CAS  Google Scholar 

  • Rojas A, Duque E, Mosqueda G, Golden G, Hurtado A, Ramos JL, Segura A (2001) Three efflux pumps are required to provide efficient tolerance to toluene in Pseudomonas putida DOT-T1E. J Bacteriol 183:3967–3973

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Segura A, Duque E, Rojas A et al (2004) Fatty acid biosynthesis is involved in solvent tolerance in Pseudomonas putida DOT-T1E. Environ Microbiol 6:416–423

    Article  PubMed  CAS  Google Scholar 

  • Shapiguzov A, Lyukevich AA, Allakhverdiev SI, Sergeyenko TV, Suzuki I, Murata N, Los DA (2005) Osmotic shrinkage of cells of Synechocystis sp. PCC 6803 by water efflux via aquaporins regulates osmostress-inducible gene expression. Microbiology 151:447–455

    Article  PubMed  CAS  Google Scholar 

  • Sharma NK, Pandey J, Gupta N, Jain RK (2007) Growth and physiological response of Arthrobacter protophormiae RKJ100 toward higher concentrations of o-nitrobenzoate and p-hydroxybenzoate. FEMS Microbiol Lett 271:65–70

    Article  PubMed  CAS  Google Scholar 

  • Takasaki S, Kobata A (1976) Purification and characterization of an endo-beta-galactosidase produced by Diplococcus pneumoniae. J Biol Chem 251:3603–3609

    PubMed  CAS  Google Scholar 

  • Takegawa K, Yamaguchi S, Kondo A, Iwamoto H, Nakoshi M, Kato I, Iwahara S (1991) Transglycosylation activity of endo-beta-N-acetylglucosaminidase from Arthrobacter protophormiae. Biochem Int 24:849–855

    PubMed  CAS  Google Scholar 

  • Tarentino AL, Maley F (1974) Purification and properties of an endo-beta-N-acetylglucosaminidase from Streptomyces griseus. J Biol Chem 249:811–817

    PubMed  CAS  Google Scholar 

  • Taylor M, Tuffin M, Burton S, Eley K, Cowan D (2008) Microbial responses to solvent and alcohol stress. Biotechnol J 3:1388–1397

    Article  PubMed  CAS  Google Scholar 

  • Wackett LP (1997) Bacterial biocatalysis: stealing a page from nature’s book. Nat Biotechnol 15:415–416

    Article  PubMed  CAS  Google Scholar 

  • Wang LX (2008) Chemoenzymatic synthesis of glycopeptides and glycoproteins through endoglycosidase-catalyzed transglycosylation. Carbohydr Res 343:1509–1522

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Yin J, Li L, Shaw N et al (2009) Structural basis and catalytic mechanism for the dual functional endo-beta-N-acetylglucosaminidase A. PLoS ONE 4:e4658

    Article  PubMed Central  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This study was partly supported by Council for Scientific and Industrial Research (CSIR) India and Department of Biotechnology (DBT) India. JP and FK acknowledge the research fellowships awarded by CSIR. We are thankful to John Oakeshott and Rinku Pandey for critically reading the manuscript. All other authors also wish to acknowledge the inspiration of RKJ, who fell ill during the course of this study and passed before the manuscript was ready for communication.

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  1. Janmejay Pandey

    Present address: Department of Biotechnology, Central University of Rajasthan, NH-8, Bandarsindri, Ajmer, 305801, Rajasthan, India

Authors and Affiliations

  1. Institute of Microbial Technology, Sector 39-A, Chandigarh, 160036, India

    Janmejay Pandey, Fazlurrahman Khan, Vivek Mahajan, Mahima Pant, Rakesh K. Jain & Gunjan Pandey

  2. CSIRO Ecosystem Sciences, Clunies Ross Street, Acton, ACT, 2601, Australia

    Gunjan Pandey

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Correspondence to Janmejay Pandey.

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Communicated by A. Driessen.

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Pandey, J., Khan, F., Mahajan, V. et al. Evidence for vital role of endo-β-N-acetylglucosaminidase in the resistance of Arthrobacter protophormiae RKJ100 towards elevated concentrations of o-nitrobenzoate. Extremophiles 18, 491–500 (2014). https://doi.org/10.1007/s00792-014-0632-2

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  • DOI: https://doi.org/10.1007/s00792-014-0632-2

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