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Two unusual chlorocatechol catabolic gene clusters in Sphingomonas sp. TFD44

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Abstract

The genes responsible for the degradation of 2,4-dichlorophenoxyacetate (2,4-D) by α-Proteobacteria have previously been difficult to detect by using gene probes or polymerase chain reaction (PCR) primers. PCR products of the chlorocatechol 1,2-dioxygenase gene, tfdC, now allowed cloning of two chlorocatechol gene clusters from the Sphingomonas sp. strain TFD44. Sequence characterization showed that the first cluster, tfdD,RFCE, comprises all the genes necessary for the conversion of 3,5-dichlorocatechol to 3-oxoadipate, including a presumed regulatory gene, tfdR, of the LysR-type family. The second gene cluster, tfdC2E2F2, is incomplete and appears to lack a chloromuconate cycloisomerase gene and a regulatory gene. Purification and N-terminal sequencing of selected enzymes suggests that at least representatives of both gene clusters (TfdD of cluster 1 and TfdC2 of cluster 2) are induced during the growth of strain TFD44 with 2,4-D. A mutant constructed to contain an insertion in the chloromuconate cycloisomerase gene tfdD still was able to grow with 2,4-D, but more slowly and with a longer lag phase. This, and the detection of additional activity peaks during protein purification suggest that strain TFD44 harbors at least another chloromuconate cycloisomerase gene. The sequence of the tfdCE region was almost identical to that of a partially characterized chlorocatechol catabolic gene cluster of Sphingomonas herbicidovorans MH, whereas the sequence of the tfdC2E2F2 cluster was different. The similarity of the predicted proteins of the tfdD,RFCE and tfdC2E2F2 clusters to known sequences of other Proteobacteria in the database ranged from 42 to 61% identical positions for the first cluster and from 45.5 to 58% identical positions for the second cluster. Between both clusters, the similarities of their predicted proteins ranged from 44.5 to 64% identical positions. Thus, both clusters (together with those of S. herbicidovorans MH) represent deep-branching lines in the respective dendrograms, and the sequence information will help future primer design for the detection of corresponding genes in the environment.

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Acknowledgements

The investigations reported here were funded by grant 0311770 from the German Federal Ministry of Education and Research. We thank Hans-Joachim Knackmuss for the opportunity to perform the initial part of the experiments in the Institute for Microbiology at the University of Stuttgart. We thank James M. Tiedje for providing strain TFD44 and Walter Reineke for chloromuconic acids.

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  1. Interdisciplinary Ecological Center, Technische Universität Bergakademie Freiberg, Leipziger Straße 29, 09599, Freiberg, Germany

    Monika Thiel, Stefan R. Kaschabek, Janosch Gröning, Margit Mau & Michael Schlömann

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  2. Stefan R. Kaschabek
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  3. Janosch Gröning
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  4. Margit Mau
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  5. Michael Schlömann
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Correspondence to Michael Schlömann.

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Thiel, M., Kaschabek, S.R., Gröning, J. et al. Two unusual chlorocatechol catabolic gene clusters in Sphingomonas sp. TFD44. Arch Microbiol 183, 80–94 (2005). https://doi.org/10.1007/s00203-004-0748-3

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  • DOI: https://doi.org/10.1007/s00203-004-0748-3

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