Indene bioconversion by a toluene inducible dioxygenase of Rhodococcus sp. I24

Horst Priefert, Xian M. O'Brien, Philip A. Lessard, Annette F. Dexter, Ellen E. Choi, Sladjana Tomic, Geeta Nagpal, Jennie J. Cho, Melina Agosto, Lucy Yang, Sheri L. Treadway, Lance Tamashiro, Matthew Wallace, Anthony J. Sinskey*

*Corresponding author for this work

Research output: Contribution to journalArticle

22 Scopus citations

Abstract

Rhodococcus sp. I24 can oxygenate indene via at least three independent enzyme activities: (i) a naphthalene inducible monooxygenase (ii) a naphthalene inducible dioxygenase, and (iii) a toluene inducible dioxygenase (TID). Pulsed field gel analysis revealed that the I24 strain harbors two megaplasmids of ∼340 and ∼50 kb. Rhodococcus sp. KY1, a derivative of the I24 strain, lacks the ∼340 kb element as well as the TID activity. Southern blotting and sequence analysis of an indigogenic, I24-derived cosmid suggested that an operon encoding a TID resides on the ∼340 kb element. Expression of the tid operon was induced by toluene but not by naphthalene. In contrast, naphthalene did induce expression of the nid operon, encoding the naphthalene dioxygenase in 124. Cell free protein extracts of Escherichia coli cells expressing tidABCD were used in HPLC-based enzyme assays to characterize the indene bioconversion of TID in vitro. In addition to 1-indenol, indene was transformed to cis-indandiol with an enantiomeric excess of 45.2% of cis-(1S,2R)-indandiol over cis-(1R,2S)-indandiol, as revealed by chiral HPLC analysis. The Km of TID for indene was 380 μM. The enzyme also dioxygenated naphthalene to cis-dihydronaphthalenediol with an activity of 78% compared to the formation of cis-indandiol from indene. The Km of TID for naphthalene was 28 μM. TID converted only trace amounts of toluene to 1,2-dihydro-3- methylcatechol after prolonged incubation time. The results indicate the role of the tid operon in the bioconversion of indene to 1-indenol and cis-(1S,2R)-indandiol by Rhodococcus sp. I24.

Original languageEnglish (US)
Pages (from-to)168-176
Number of pages9
JournalApplied Microbiology and Biotechnology
Volume65
Issue number2
DOIs
StatePublished - Aug 2004

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology

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    Priefert, H., O'Brien, X. M., Lessard, P. A., Dexter, A. F., Choi, E. E., Tomic, S., Nagpal, G., Cho, J. J., Agosto, M., Yang, L., Treadway, S. L., Tamashiro, L., Wallace, M., & Sinskey, A. J. (2004). Indene bioconversion by a toluene inducible dioxygenase of Rhodococcus sp. I24. Applied Microbiology and Biotechnology, 65(2), 168-176. https://doi.org/10.1007/s00253-004-1589-3