Cloning and sequencing of the gene encoding the enzyme for the reductive cleavage of diaryl ether bonds of 2,3,7,8-tetrachlorodibenzo-p-dioxin in Geobacillus thermodenitrificans UZO 3

• Published in: Journal of bioscience and bioengineering 2018-10, Vol.126 (4), p.488-496
• Main Authors: Suzuki, Yuzo, Nakamura, Masaya, Otsuka, Yuichiro, Suzuki, Nao, Ohyama, Keisuke, Kawakami, Takeshi, Sato-Izawa, Kanna, Navarro, Ronald R., Hishiyama, Shojiro, Inoue, Kouya, Kameyama, Toshiji, Takahashi, Atsushi, Katayama, Yoshihiro
• Format: Article
• Language: English
• Subjects: 2,3,7,8-Tetrachlorodibenzo-p-dioxin; Acyltransferases - chemistry; Acyltransferases - genetics; Acyltransferases - metabolism; Amino Acid Sequence; Animals; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Binding Sites; Bioremediation; Cloning; Cloning, Molecular; Degradation gene; Dioxin; Escherichia coli - genetics; Escherichia coli - metabolism; Ether - chemistry; Ether - metabolism; Geobacillus - chemistry; Geobacillus - enzymology; Geobacillus - genetics; Polychlorinated Dibenzodioxins - chemistry; Polychlorinated Dibenzodioxins - metabolism
• ISSN: 1389-1723; 1347-4421
• DOI: 10.1016/j.jbiosc.2018.04.013

We have previously reported that a cell-free extract prepared from Geobacillus thermodenitrificans UZO 3 reductively cleaves diaryl ether bonds of 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD), a dioxin with the highest toxicity, in a sequential fashion producing 3′,4′,4,5-tetrachloro-2-hydroxydiphenyl ether (TCDE) as the intermediate, and 3,4-dichlorophenol (DCP) as the final reaction product. The detection of TCDE implicated the discovery of an unprecedented dioxin-degrading enzyme that reductively cleaves the diaryl ether bonds. In this study, we report the cloning and sequencing of the dioxin reductive etherase gene dreE which codes for the 2,3,7,8-TCDD-degrading enzyme. We showed that dreE was expressed in Escherichia coli and that the product of the expression could reductively cleave diaryl ether bonds of 2,3,7,8-TCDD to produce TCDE. Furthermore, we established that the amino acid sequence encoded by dreE was homologous to an enzyme with yet unknown function that is encoded by a gene located in the riboflavin (vitamin B2) biosynthesis operon in Bacillus subtilis. We also showed that the amino acid sequence possesses a coenzyme A (CoA) binding site that is conserved in the N-acyltransferase superfamily. For the first time, the degradation of 2,3,7,8-TCDD at the molecular level using a enzyme of bacterial origin has been demonstrated. A novel mechanism model for the reductive cleavage of diaryl ether bond of 2,3,7,8-TCDD was also proposed.