Skip to main content
SpringerLink
Log in

Xenoestrogenic short ethoxy chain nonylphenol is oxidized by a flavoprotein alcohol dehydrogenase from Ensifer sp. strain AS08

  • Applied Microbial and Cell Physiology
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

The ethoxy chains of short ethoxy chain nonylphenol (NPEOav2.0, containing average 2.0 ethoxy units) were dehydrogenated by cell-free extracts from Ensifer sp. strain AS08 grown on a basal medium supplemented with NPEOav2.0. The reaction was coupled with the reduction in 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide and phenazine methosulfate. The enzyme (NPEOav2.0 dehydrogenase; NPEO-DH) was purified to homogeneity with a yield of 20% and a 56-fold increase in specific activity. The molecular mass of the native enzyme was 120 kDa, consisting of two identical monomer units (60 kDa). The gene encoding NPEO-DH was cloned, which consisted of 1,659 bp, corresponding to a protein of 553 amino acid residues. The deduced amino acid sequence agreed with the N-terminal amino acid sequence of the purified NPEO-DH. The presence of a flavin adenine dinucleotide (FAD)-binding motif and glucose–methanol–choline (GMC) oxidoreductase signature motifs strongly suggested that the enzyme belongs to the GMC oxidoreductase family. The protein exhibited homology (40–45% identity) with several polyethylene glycol dehydrogenases (PEG-DHs) of this family, but the identity was lower than those (approximately 58%) among known PEG-DHs. The substrate-binding domain was more hydrophobic compared with those of glucose oxidase and PEG-DHs. The recombinant protein had the same molecular mass as the purified NPEO-DH and dehydrogenated PEG400-2000, NPEOav2.0 and its components, and NPEOav10, but only slight or no activity was found using diethylene glycol, triethylene glycol, and PEG200.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Ahel M, Giger W (1985) Determination of alkylphenols and alkylphenol mono-and diethoxylates in environmental samples by high-performance liquid chromatography. Anal Chem 57:1577–1583

    Article  CAS  Google Scholar 

  • Ahel M, Conrad T, Giger W (1987) Persistent organic chemicals in sewage effluents. 3. Determinations of nonylphenoxy carboxylic acids by high-resolution gas chromatography/mass spectrometry and high performance liquid chromatography. Environ Sci Technol 21:697–703

    Article  CAS  PubMed  Google Scholar 

  • Brunner PH, Capri A, Marcomini A, Giger W (1988) Occurrence and behaviour of linear alkylbenze-sulphonates, nonylphenol, nonylphenol mono- and di-ethoxylates in sewage and sewage sludge treatment. Water Res 22:1465–1472

    Article  CAS  Google Scholar 

  • Cavener DR (1992) GMC oxidoreductase. A newly defined family of homologous proteins with diverse catalytic activities. J Mol Biol 223:811–814

    Article  CAS  PubMed  Google Scholar 

  • Ejlertsson J, Nilsson ML, Kylin H, Bergman A, Karlson I, Oquist M, Svensson BH (1999) Anaerobic degradation of nonylphenol mono- and di-ethoxylates in digestor sludge, landfill municipal solid waste and landfill sludge. Environ Sci Technol 33:301–306

    Article  CAS  Google Scholar 

  • Enokibara S, Kawai F (1997) Purification and characterization of an ether-cleaving enzyme involved in the metabolism of polyethylene glycol. J Ferment Bioeng 83:549–554

    Article  CAS  Google Scholar 

  • Giger W, Brunner PH, Schaffner C (1984) 4-Nonylphenol in sewage sludge: accumulation of toxic metabolites from non-ionic surfactants. Science 225:623–625

    Article  CAS  PubMed  Google Scholar 

  • Hirokawa, T, Boon-Chieng S, Mitaku S (1998) SOSUI: classification and secondary structure prediction system for membrane proteins. Bioinformatics 14:378–379

    Article  CAS  PubMed  Google Scholar 

  • John DM, White GF (1998) Mechanism for biotransformation of nonylphenol polyethoxylates to xenoestrogens in Pseudomonas putida. J Bacteriol 180:4332–4338

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kawai F (2002) Microbial degradation of polyethers. Appl Microbiol Biotechnol 58:30–38

    Article  CAS  PubMed  Google Scholar 

  • Kawai F, Kimura T, Fukaya M, Tani Y, Ogata K, Ueno T, Fukami H (1978) Bacterial oxidation of polyethylene glycol. Appl Environ Microbiol 35:679−684

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kawai F, Kimura T, Tani Y, Tamada H, Kurachi M (1980) Purification and characterization of polyethylene glycol dehydrogenase involved in the bacterial metabolism of polyethylene glycol. Appl Environ Microbiol 40:701−705

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kiess M, Hecht HJ, Kalisz HM (1998) Glucose oxidase from Penicillium amagasakiense: primary structure and comparison with other glucose–methanol–choline (GMC) oxidoreductases. Eur J Biochem 252:90–99

    Article  CAS  PubMed  Google Scholar 

  • Kobayashi T, Takahashi Y, Ito K (2001) Identification of a segment of DsbB essential for its respiration-coupled oxidation. Mol Microbiol 39:158–165

    Article  CAS  PubMed  Google Scholar 

  • Kyte J, Doolittle RF (1982) A simple method for displaying the hydropathic character of a protein. J Mol Biol 157:105–132

    CAS  PubMed  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  CAS  PubMed  Google Scholar 

  • Liu X, Tani A, Kimbara K, Kawai F (2006) Metabolic pathway of xenoestrogenic short ethoxy chain-nonylphenol to nonylphenol by aerobic bacteria, Ensifer sp. strain AS08 and Pseudomonas sp. strain AS90. Appl Microbiol Biotechnol (in press)

  • Maeda S, Mikami I (1988) Degradation of the non-ionic surfactant (polyoxyethylene-p-nonylphenyl ether) by Pseudomonas species. J Water Waste 30:1056–1063

    CAS  Google Scholar 

  • Maki H, Masuda N, Fujiwara Y, Ike M, Fujita M (1994) Degradation of alkyl phenol ethoxylates by Pseudomonas sp. strain TR01. Appl Environ Microbiol 60:2265–2271

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ochman H, Gerber AS, Hart DL (1988) Genetic applications of an inverse polymerase chain reaction. Genetics 120:621–623

    CAS  PubMed  PubMed Central  Google Scholar 

  • Ohta T, Kawabata T, Nishikawa K, Tani A, Kimbara K, Kawai F (2006) Analysis of amino acid residues involved in the catalysis of polyethylene glycol dehydrogenase from Sphingopyxis terrae using kinetic characterization of mutants based on 3D molecular modeling. Appl Environ Microbiol 72:4388–4396

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

    Google Scholar 

  • Sugimoto M, Tanabe M, Hataya M, Enokibara S, Duine JA, Kawai F (2001) The first step in polyethylene glycol degradation by sphingomonads proceeds via a flavoprotein alcohol dehydrogenase containing flavin adenine dinucleotide. J Bacteriol 183:6694–6698

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Szeto TH, Rowland SL, Rothfield LI, King GF (2002) Membrane localization of MinD is mediated by a C-terminal motif that is conserved across eubacteria, archaea, and chloroplasts. Proc Natl Acad Sci U S A 99:15693–15698

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wohlfahrt G, Witt S, Hendle J, Schomburg D, Kalisz HM, Hecht HJ (1999) 1.8 and 1.9 Å resolution structures of the Penicillium amagasakiense and Aspergillus niger glucose oxidases as a basis for modelling substrate complexes. Acta Crystallogr D55:969–977

    CAS  Google Scholar 

Download references

Acknowledgments

This work was partly supported by the Research Grant for Encouragement of Studies to F.K. and X.L. from the Graduate School of Natural Science and Technology, Okayama University.

Author information

Authors and Affiliations

  1. Laboratory of Applied Microbiology, Research Institute for Bioresources, Okayama University, Kurashiki, 2-20-1 Chuo, Kurashiki, 710-0046, Japan

    Xin Liu, Akio Tani, Kazuhide Kimbara & Fusako Kawai

Authors
  1. Xin Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Akio Tani
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kazuhide Kimbara
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Fusako Kawai
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Fusako Kawai.

Additional information

English edition: The paper was edited by a native speaker through American Journal Experts (http://www.journalexperts.com).

Rights and permissions

Reprints and permissions

About this article

Cite this article

Liu, X., Tani, A., Kimbara, K. et al. Xenoestrogenic short ethoxy chain nonylphenol is oxidized by a flavoprotein alcohol dehydrogenase from Ensifer sp. strain AS08. Appl Microbiol Biotechnol 73, 1414–1422 (2007). https://doi.org/10.1007/s00253-006-0620-2

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-006-0620-2

Keywords

Search

Navigation