Mechanism controlling the extended lag period associated with vinyl chloride starvation in Nocardioides sp. strain JS614

Abstract

The extended lag period associated with vinyl chloride (VC) starvation in VC- and ethene-assimilating Nocardioides sp. strain JS614 was examined. The extended lag periods were variable (3-7 days), only associated with growth on VC or ethene, and were observed in VC- or ethene-grown cultures following 24 h carbon starvation and mid-exponential phase cultures grown on non-alkene carbon sources (e.g. acetate). Alkene monooxygenase (AkMO) and epoxyalkane:coenzyme M transferase (EaCoMT) are the initial enzymes of VC and ethene biodegradation in strain JS614. Reverse-transcription PCR confirmed that the AkMO gene etnC was expressed in response to epoxyethane, a metabolic intermediate of ethene biodegradation. Epoxyethane (0.5 mM) eliminated the extended lag period in both starved and mid-exponential phase cultures, suggesting that epoxyethane accumulation activates AkMO expression in strain JS614. AkMO activity in ethene-grown cultures was not detected after 6.7 h of carbon starvation, while 40% of the initial EaCoMT activity remained after 24 h. Acetate eliminated the extended lag period in starved cultures but not in mid-exponential phase cultures suggesting that acetate reactivates extant AkMO in starved VC- or ethene-grown cultures. The imbalance between AkMO and EaCoMT activities during starvation likely contributes to the extended lag period by delaying epoxide accumulation and subsequent AkMO induction.

Identification of Polypeptides Expressed in Response to Vinyl Chloride, Ethene, and Epoxyethane in Nocardioides sp. Strain JS614 by Using Peptide Mass Fingerprinting

“…3) that contained known VC/ethene biodegradation genes (18), suggesting that they are likely to participate in VC/ethene metabolism. Identification of AkMO subunit (EtnC), EaCoMT (EtnE), putative CoM biosynthesis protein ComA, and a probable CoA transferase in cell extracts from VC-and ethenegrown cells is consistent with previous reports (17,18), confirming the robustness of this proteomics approach. Five proteins not previously reported to be involved in VC/ethene metabolism were also identified.…”

“…Reverse transcription (RT)-PCR assays were conducted to provide independent confirmation of proteomics results. Total RNA was extracted and purified, and RT-PCR was performed as previously described (17,18), using 0.1 ng RNA template. JS614 genomic DNA, extracted as described previously (3), was used as the positive RT-PCR control.…”

Aerobic biodegradation of the chloroethenes: pathways, enzymes, ecology, and evolution

“…However, because VC and ethene would be converted to epoxides under the experimental conditions used in the JS60 study, this experiment did not allow the actual inducer to be pinpointed. Later experiments with Nocardioides strain JS614 addressed this, firstly by showing by RT‐PCR that etnC was transcribed in epoxyethane‐grown cultures, and secondly, by showing that epoxyethane addition allowed starved JS614 cells to rapidly resume alkene degradation (Mattes et al ., 2007). The JS614 data therefore indicate that epoxyalkanes are the inducers of the VC/ethene catabolic pathway.…”

Characterization of Microbes Capable of Using Vinyl Chloride and Ethene as Sole Carbon and Energy Sources by Anaerobic Oxidation

“…Other organisms capable of growing on VC have been found to show a lag in growth (Verce et al, 2000). For some isolates, this lag has been attributed to the absence of VC-epoxide in cells needed to induce alkene monooxygenase (Mattes et al, 2007).…”