• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of aemPermissionsJournals.ASM.orgJournalAEM ArticleJournal InfoAuthorsReviewers
Appl Environ Microbiol. Dec 1985; 50(6): 1409–1413.
PMCID: PMC238772

Use of cloned genes of Pseudomonas TOL plasmid to effect biotransformation of benzoates to cis-dihydrodiols and catechols by Escherichia coli cells.


DNA fragments containing the xylD and xylL genes, which specify the broad-specificity enzymes toluate-1,2-dioxygenase and 3,5-cyclohexadiene-1,2-diol-1-carboxylic acid dehydrogenase, respectively, of TOL plasmid pWW0-161 of Pseudomonas putida have previously been cloned in the pBR322 vector plasmid (P.R. Lehrbach, J. Zeyer, W. Reinecke, H.-J. Knackmuss, and K. N. Timmis, J. Bacteriol. 158:1025-1032, 1984). In this study, Escherichia coli cells containing hybrid plasmids carrying the cloned xylD or xylDL genes quantitatively transformed 14C-ring- and 14C-carboxy-labeled benzoate to the pathway intermediates 3,5-cyclohexadiene-1,2-diol-1-carboxylic acid (cis-dihydrodiol) and catechol, respectively. Like P. putida cells, E. coli cells containing the xylD gene transformed a variety of chloro- and hydrocarbon-substituted benzoates. The toluate-1,2-dioxygenase produced in E. coli thus exhibited the broad-substrate-specificity properties of the enzyme in P. putida. Turnover rates by the enzymes in these two bacteria are compared.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.0M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Dorn E, Knackmuss HJ. Chemical structure and biodegradability of halogenated aromatic compounds. Substituent effects on 1,2-dioxygenation of catechol. Biochem J. 1978 Jul 15;174(1):85–94. [PMC free article] [PubMed]
  • Ensley BD, Ratzkin BJ, Osslund TD, Simon MJ, Wackett LP, Gibson DT. Expression of naphthalene oxidation genes in Escherichia coli results in the biosynthesis of indigo. Science. 1983 Oct 14;222(4620):167–169. [PubMed]
  • Franklin FC, Bagdasarian M, Bagdasarian MM, Timmis KN. Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7458–7462. [PMC free article] [PubMed]
  • Franklin FC, Lehrbach PR, Lurz R, Rueckert B, Bagdasarian M, Timmis KN. Localization and functional analysis of transposon mutations in regulatory genes of the TOL catabolic pathway. J Bacteriol. 1983 May;154(2):676–685. [PMC free article] [PubMed]
  • Harayama S, Lehrbach PR, Timmis KN. Transposon mutagenesis analysis of meta-cleavage pathway operon genes of the TOL plasmid of Pseudomonas putida mt-2. J Bacteriol. 1984 Oct;160(1):251–255. [PMC free article] [PubMed]
  • Inouye S, Nakazawa A, Nakazawa T. Molecular cloning of gene xylS of the TOL plasmid: evidence for positive regulation of the xylDEGF operon by xylS. J Bacteriol. 1981 Nov;148(2):413–418. [PMC free article] [PubMed]
  • Kunz DA, Chapman PJ. Catabolism of pseudocumene and 3-ethyltoluene by Pseudomonas putida (arvilla) mt-2: evidence for new functions of the TOL (pWWO) plasmid. J Bacteriol. 1981 Apr;146(1):179–191. [PMC free article] [PubMed]
  • Lehrbach PR, Timmis KN. Genetic analysis and manipulation of catabolic pathways in Pseudomonas. Biochem Soc Symp. 1983;48:191–219. [PubMed]
  • Lehrbach PR, Zeyer J, Reineke W, Knackmuss HJ, Timmis KN. Enzyme recruitment in vitro: use of cloned genes to extend the range of haloaromatics degraded by Pseudomonas sp. strain B13. J Bacteriol. 1984 Jun;158(3):1025–1032. [PMC free article] [PubMed]
  • Mermod N, Lehrbach PR, Reineke W, Timmis KN. Transcription of the TOL plasmid toluate catabolic pathway operon of Pseudomonas putida is determined by a pair of co-ordinately and positively regulated overlapping promoters. EMBO J. 1984 Nov;3(11):2461–2466. [PMC free article] [PubMed]
  • Murray K, Duggleby CJ, Sala-Trepat JM, Williams PA. The metabolism of benzoate and methylbenzoates via the meta-cleavage pathway by Pseudomonas arvilla mt-2. Eur J Biochem. 1972 Jul 24;28(3):301–310. [PubMed]
  • Reineke W, Knackmuss HJ. Chemical structure and biodegradability of halogenate aromatic compounds. Substituent effects on 1,2-dioxygenation of benzoic acid. Biochim Biophys Acta. 1978 Sep 6;542(3):412–423. [PubMed]
  • Reiner AM. Metabolism of aromatic compounds in bacteria. Purification and properties of the catechol-forming enzyme, 3,5-cyclohexadiene-1,2-diol-1-carboxylic acid (NAD + ) oxidoreductase (decarboxylating). J Biol Chem. 1972 Aug 25;247(16):4960–4965. [PubMed]
  • Reiner AM, Hegeman GD. Metabolism of benzoic acid by bacteria. Accumulation of (-)-3,5-cyclohexadiene-1,2-diol-1-carboxylic acid by mutant strain of Alcaligenes eutrophus. Biochemistry. 1971 Jun 22;10(13):2530–2536. [PubMed]
  • Williams PA, Murray K. Metabolism of benzoate and the methylbenzoates by Pseudomonas putida (arvilla) mt-2: evidence for the existence of a TOL plasmid. J Bacteriol. 1974 Oct;120(1):416–423. [PMC free article] [PubMed]
  • Worsey MJ, Williams PA. Metabolism of toluene and xylenes by Pseudomonas (putida (arvilla) mt-2: evidence for a new function of the TOL plasmid. J Bacteriol. 1975 Oct;124(1):7–13. [PMC free article] [PubMed]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)


Cited by other articles in PMC

See all...


Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...