Archives of Microbiology

, Volume 188, Issue 1, pp 55–68 | Cite as

Degradation of aromatic compounds by Acinetobacter radioresistens S13: growth characteristics on single substrates and mixtures

  • Roberto MazzoliEmail author
  • Enrica Pessione
  • Maria G. Giuffrida
  • Paolo Fattori
  • Cristina Barello
  • Carlo Giunta
  • Nicholas D. Lindley
Original Paper


Acinetobacter radioresistens S13 is able to grow on phenol or benzoate as the sole carbon and energy source: both these compounds are catabolized through the β-ketoadipate pathway. Genes encoding the catabolic enzymes for degradation of aromatic compounds are localized on A. radioresistens S13 chromosome and organized in, at least, two distinct sets, one for benzoate degradation and another for phenol catabolism. In the present study, the growth and biodegradation kinetics for benzoate and phenol, and an easily metabolized substrate (acetate) were established. Benzoate was degraded slower and supports a less rapid and efficient growth than either acetate or phenol. A combined transcript-proteomic analysis of some of the major catabolic genes and their products nonetheless has shown that benzoate induces the expression of both benzoate and phenol catabolic operons. This result was confirmed by the fact that benzoate-acclimatized bacteria were rapidly able to degrade phenol too. Finally, the growth and biodegradation kinetics for different mixtures of acetate, benzoate and phenol were determined. Results indicate that a hierarchy of substrate utilization, benzoate > acetate > phenol, occurred: benzoate was the preferred substrate, despite its lower growth and biodegradation parameters. Hypotheses explaining these unusual metabolic features of A. radioresistens S13 are discussed.


Proteome Transcript analysis Ortho cleavage pathway Benzoate dioxygenase Phenol hydroxylase 



Alkaline phosphatase


Phenol hydroxylase and its components (PHO oxygenase, PHR reductase, PHI intermediate)


Benzoate dioxygenase and its components (BDR reductase, BDO oxygenase)


Dihydroxybenzoate dehydrogenase

C12O IsoA (gene catAA)

Catechol 1,2-dioxygenase isoenzyme A

C12O IsoB (gene catAB)

Catechol 1,2-dioxygenase isoenzyme B

MCI (gene catB)

Muconate cycloisomerase

MLI (gene catC)

Muconolactone isomerase


β-ketoadipil-CoA thiolase


The gene encoding BDOα


The gene encoding PHOα


Bacterial specific growth rate


Substrate consumption rate


Bacterial growth yield


Millimoles of carbon



We would like to thank Dr. Sergine Even for invaluable aid in the optimization of transcript techniques. This study received financial support from FIRB. Roberto Mazzoli received a European Union Marie Curie Training Site grant (HPMT-2000-00135) while travel expenses were possible due to financial support from the Franco-Italian Galileo bilateral support scheme.

Supplementary material

203_2007_223_MOESM1_ESM.ppt (155 kb)
ESM1 (PPT 155 KB)


  1. Ampe F (1995) PhD Thesis, INSA-Toulouse, ToulouseGoogle Scholar
  2. Ampe F, Lindley ND (1995) Acetate utilization is inhibited by benzoate in Alcaligenes eutrophus: evidence for transcriptional control of the expression of acoE coding for acetyl coenzyme-A synthetase. J Bacteriol 177:5826–5833PubMedGoogle Scholar
  3. Ampe F, Lindley ND (1996) Flux limitations in the ortho pathway of benzoate degradation of Alcaligenes eutrophus: metabolite overflow and induction of the meta pathway at high substrate concentrations. Microbiology 142:1807–1817PubMedCrossRefGoogle Scholar
  4. Ampe F, Leonard D, Lindley ND (1996) Growth performance and flux determine substrate preference of Alcaligenes eutrophus during growth on acetate plus aromatic compound mixtures. Appl Microbiol Biotechnol 46:562–569CrossRefGoogle Scholar
  5. Ampe F, Uribelarrea JL, Aragao GMF, Lindley ND (1997) Benzoate degradation via the ortho pathway in Alcaligenes eutrophus is perturbed by succinate. Appl Environ Microbiol 63:2765–2770PubMedGoogle Scholar
  6. Ampe F, Leonard D, Lindley ND (1998) Repression of phenol catabolism by organic acids in Ralstonia eutropha. Appl Environ Microbiol 64:1–6PubMedGoogle Scholar
  7. Brzostowicz PC, Reams AB, Clark TJ, Neidle EL (2003) Transcriptional cross-regulation of the catechol and protocatechuate branches of the β-ketoadipate pathway contributes to carbon source-dependent expression of the Acinetobacter sp. strain ADP1 pobA gene. Appl Environ Microbiol 60:1598–1606CrossRefGoogle Scholar
  8. Caposio P, Pessione E, Giuffrida MG, Conti A, Landolfo S, Giunta C, Gribaudo G (2002) Cloning and characterization of two catechol 1,2-dioxygenase genes from Acinetobacter radioresistens S13. Res Microbiol 153:69–74PubMedCrossRefGoogle Scholar
  9. Cases I, de Lorenzo V (2005) Promoters in the environment: transcriptional regulation in its natural context. Nat Rev Microbiol 3:105–118PubMedCrossRefGoogle Scholar
  10. Clark TJ, Momany C, Neidle EL (2002) The benPK operon, proposed to play a role in transport, is part of a regulon for benzoate catabolism in Acinetobacter sp. Strain ADPI. Microbiology 148:1213–1223PubMedGoogle Scholar
  11. Collier DN, Hager PW, Phibbs PV Jr (1996) Catabolite repression control in the Pseudomonads. Res Microbiol 147:551–561PubMedCrossRefGoogle Scholar
  12. Dal S, Steiner I, Gerischer U (2002) Multiple operons connected with catabolism of aromatic compounds in Acinetobacter sp. strain ADP1 are under carbon catabolite repression. J Mol Microbiol Biotechnol 4:389–404PubMedGoogle Scholar
  13. Divari S, Valetti F, Caposio P, Pessione E, Cavaletto M, Griva E, Gribaudo G, Gilardi G, Giunta C (2003) The oxygenase component of phenol hydroxylase from Acinetobacter radioresistens S13. Eur J Biochem 270:2244–2253PubMedCrossRefGoogle Scholar
  14. Fontaine L, Even S, Soucaille P, Lindley ND, Cocaign-Bousquet M (2001) Transcript quantification based on chemical labeling of RNA associated with fluorescent detection. Anal Biochem 298:246–252PubMedCrossRefGoogle Scholar
  15. Gaines GL III, Smith L, Neidle EL (1996) Novel nuclear magnetic resonance spectroscopy methods demonstrate preferential carbon source utilization by Acinetobacter calcoaceticus. J Bacteriol 178:6833–6841PubMedGoogle Scholar
  16. Giuffrida MG, Pessione E, Mazzoli R, Dellavalle G, Barello C, Conti A, Giunta C (2001) Media containing aromatic compounds induce peculiar proteins in Acinetobacter radioresistens, as revealed by proteome analysis. Electrophoresis 22:1705–1711PubMedCrossRefGoogle Scholar
  17. Griva E, Pessione E, Divari S, Valetti F, Cavaletto M, Rossi GL, Giunta C (2003) Phenol hydroxylase from Acinetobacter radioresistens S13. Isolation and characterization of the regulatory component. Eur J Biochem 270:1434–1440PubMedCrossRefGoogle Scholar
  18. Harayama S (1991) Induction kinetics of RNA and proteins in exponentially growing organisms. Biochem Biophys Res Commun 180:913–919PubMedCrossRefGoogle Scholar
  19. Heinaru E, Viggor S, Vedler E, Truu J, Merimaa M, Heinaru A (2001) Reversible accumulation of p-hydroxybenzoate and catechol determines the sequential decomposition of phenolic compounds in mixed substrate cultivations in pseudomonads. FEMS Microbiol Ecol 37:79–89CrossRefGoogle Scholar
  20. Juni E (1978) Genetics and physiology of Acinetobacter. Annu Rev Microbiol 32:349–371PubMedCrossRefGoogle Scholar
  21. Kim SI, Song SY, Kim KW, Ho EM, Oh KH (2003) Proteomic analysis of the benzoate degradation pathway in Acinetobacter sp. KS-1. Res Microbiol 154:697–703PubMedCrossRefGoogle Scholar
  22. Lallai A, Mura G (1989) pH variations during phenol biodegradation in mixed cultures of microorganisms. Water Res 23:1335–1338CrossRefGoogle Scholar
  23. Lallai A, Mura G, Miliddi R, Mastinu C (1988) Effect of pH on growth of mixed cultures in batch reactor. Biotechnol Bioeng 31:130–134CrossRefPubMedGoogle Scholar
  24. Lee PS, Shaw LB, Choe LH, Mehra A, Hatzimanikatis V, Lee KH (2003) Insights into the relation between mRNA and protein expression patterns: II. Experimental observations in Escherichia coli. Biotechnol Bioeng 84:834–841PubMedCrossRefGoogle Scholar
  25. MacGregor CH, Wolff JA, Arora SK, Hylemon PB, Phibbs PV (1992) Catabolite repression control in Pseudomonas aeruginosa. In: Galli E, Silver S, Witholt B (eds) Pseudomonas: molecular biology and biotechnology. American Society for Microbiology, Washington DC, pp 198–206Google Scholar
  26. McFall SM, Abraham B, Narsolis CG, Chakrabarty AM (1997) A tricarboxylic acid cycle intermediate regulating transcription of a chloroaromatic biodegradative pathway: fumarate-mediated repression of the clcABD operon. J Bacteriol 179:6729–6735PubMedGoogle Scholar
  27. Mehra A, Lee KH, Hatzimanikatis V (2003) Insights into the relation between mRNA and protein expression patterns: I. Theoretical considerations. Biotechnol Bioeng 84:822–833PubMedCrossRefGoogle Scholar
  28. Messer M, Griffiths M, Rismiller PD, Shaw DC (1997) Lactose synthesis in a monotreme, the echidna (Tachyglossus aculeatus): isolation and amino acid sequence of echidna alpha-lactalbumin. Comp Biochem Physiol B Biochem Mol Biol 118:403–410PubMedCrossRefGoogle Scholar
  29. Navon-Venezia S, Zosim Z, Gottlieb A, Legmann R, Carmeli S, Ron EZ, Rosenberg E (1995) Alasan, a new bioemulsifier from Acinetobacter radioresistens. Appl Environ Microbiol 61:3240–3244PubMedGoogle Scholar
  30. Nichols NN, Harwood CS (1995) Repression of 4-hydroxybenzoate transport and degradation by benzoate: a new layer of regulatory control in the Pseudomonas putida β-ketoadipate pathway. J Bacteriol 177:7033–7040PubMedGoogle Scholar
  31. Nichols NN, Harwood CS (1997) PcaK, a high-affinity permease for the aromatic compounds 4-hydroxybenzoate and protocatechuate from Pseudomonas putida. J Bacteriol 179:5056–5061PubMedGoogle Scholar
  32. Perez-Pantoja D, Ledger T, Pieper DH, Gonzalez B (2003) Efficient turnover of chlorocatechols is essential for growth of Ralstonia eutropha JMP134(pJP4) in 3-chlorobenzoic acid. J Bacteriol 185:1534–1542PubMedCrossRefGoogle Scholar
  33. Pessione E, Giunta C (1997) Acinetobacter radioresistens metabolizing aromatic compounds. 2. Biochemical and microbiological characterization of the strain. Microbios 89:105–117PubMedGoogle Scholar
  34. Pessione E, Bosco F, Specchia V, Giunta C (1996) Acinetobacter radioresistens metabolizing aromatic compounds. 1. Optimization of the operative conditions for phenol degradation. Microbios 88:213–221PubMedGoogle Scholar
  35. Pessione E, Divari S, Griva E, Cavaletto M, Rossi GL, Gilardi G, Giunta C (1999) Phenol hydroxylase from Acinetobacter radioresistens is a multicomponent enzyme. Purification and characterization of the reductase moiety. Eur J Biochem 265:549–555PubMedCrossRefGoogle Scholar
  36. Pessione E, Giuffrida MG, Mazzoli R, Caposio P, Landolfo S, Conti A, Giunta C, Gribaudo G (2001) The catechol 1,2 dioxygenase system of Acinetobacter radioresistens: isoenzymes, inductors and gene localisation. Biol Chem 382:1253–1261PubMedCrossRefGoogle Scholar
  37. Pessione E, Giuffrida MG, Prunotto L, Barello C, Mazzoli R, Fortunato D, Conti A, Giunta C (2003) Membrane proteome of Acinetobacter radioresistens S13 during aromatic exposure. Proteomics 3:1070–1076PubMedCrossRefGoogle Scholar
  38. Rauhut R, Klug G (1999) mRNA degradation in bacteria. FEMS Microbiol Rev 23:353–370PubMedCrossRefGoogle Scholar
  39. Ross T, McMeekin TA (2003) Modeling microbial growth within food safety risk assessments. Risk Anal 23:179–197PubMedCrossRefGoogle Scholar
  40. Saier MH (1998) Multiple mechanisms controlling carbon metabolism in bacteria. Biotechnol Bioeng 58:170–174PubMedCrossRefGoogle Scholar
  41. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar
  42. Sanchez JC, Rouge V, Pisteur M, Ravier F, Tonella L, Moosmayer M, Wilkins MR, Hochstrasser DF (1997) Improved and simplified in-gel sample application using reswelling of dry immobilized pH gradients. Electrophoresis 18:324–327PubMedCrossRefGoogle Scholar
  43. Schirmer F, Hillen W (1998) The Acinetobacter calcoaceticus NCIB8250 mop operon mRNA is differentially degraded, resulting in a higher level of the 3′ CatA-encoding segment than of the 5′ phenolhydroxylase-encoding portion. Mol Gen Genet 257:330–337PubMedGoogle Scholar
  44. Sokol W, Howell JA (1981) Kinetics of phenol oxydation by washed cells. Biotechnol Bioeng 23:2039–2049CrossRefGoogle Scholar
  45. Stulke J, Hillen W (2000) Regulation of carbon catabolism in Bacillus species. Annu Rev Microbiol 54:849–880PubMedCrossRefGoogle Scholar
  46. Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Roberto Mazzoli
    • 1
    • 3
    Email author
  • Enrica Pessione
    • 1
  • Maria G. Giuffrida
    • 2
  • Paolo Fattori
    • 1
  • Cristina Barello
    • 2
  • Carlo Giunta
    • 1
  • Nicholas D. Lindley
    • 3
  1. 1.Dipartimento di Biologia Animale e dell’UomoUniversità di TorinoTorinoItaly
  2. 2.ISPA-CNRColleretto Giacosa (To)Italy
  3. 3.Laboratoire d’Ingénierie des Systèmes Biologiques et des Procédés, UMR CNRS/INRA/INSAInstitut National des Sciences AppliquéesToulouseFrance

Personalised recommendations