1,4-Dioxane (dioxane) is a groundwater contaminant of emerging concerns given its carcinogenic potential and widespread occurrence. To date, there are more than ten bacterial species that have been isolated to be capable of utilizing dioxane as the sole carbon and energy source. However, tetrahydrofuran (THF) monooxygenase is so far the only group of bacterial enzymes that have been well characterized and studied due to their critical role in initiating the oxidation of dioxane and other cyclic ethers. The gene cluster thmADBC encodes four components of the THF monooxygenase. Recent studies demonstrated thmADBC is absent in many newly isolated dioxane degraders, which represents a knowledge gap to uncover novel enzymes/genes involved in the biodegradation of dioxane. In this study, we sequenced the genome of Mycobacterium dioxanotrophicus PH-06, a bacterial strain exhibiting superior degradation kinetics and affinity towards dioxane. Whole genome sequence using PacBio RS II long-read sequencer unveiled the existence of a novel putative propane monooxygenase gene cluster (prmABCD), exhibiting a high similarity to different soluble di-iron monooxygenase genes found in (hydrocarbon-degrading) Rhodococcus wratislaviensis IFP2016 (89%) and Mycobacterium chubuense NBB4 (87%). Both mRNA sequencing and Reverse Transcription Quantitative PCR (RT-qPCR) analysis have indicated that all four components of this prmABCD gene cluster were upregulated when fed with dioxane in comparison with glucose or acetate as the negative control. Furthermore, dioxane biotransformation capability of this putative propane monooxygenase was confirmed in Mycobacterium smegmatis mc2 155 cells heterologously expressing the PH-06 prmABCD gene cluster. The expression clones harboring prmABCD sustain their activity to degrade dioxane and THF after induction, revealing their essential role in the oxidation of cyclic ethers. Identification of this novel enzyme is of great significance to advance our understanding of dioxane metabolic pathways and develop molecular biomarkers to assess bioremediation potential at contaminated sites.

Uncovering a novel bacterial monooxygenase that breaks down 1,4-dioxane

256th National Meeting (2018)

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