Past operational practices at chemical manufacturing facilities and widespread use of synthetic chemicals in agriculture, industry, and military operations have introduced many anthropogenic compounds to the biosphere. Some of them are readily biodegradable as a likely consequence of bacterial evolution of efficient degradation pathways, whereas others are partially degraded or persistent in the environment. Insight about biodegradation mechanisms and distribution of bacteria responsible provide the basis to predict the fate of synthetic chemicals in the environment and to enable bioremediation.
The main focus of the research described here encompasses basic science to discover pathways and evolutionary implications of aerobic biodegradation of two specific synthetic chemicals, cis-dichloroethene (cDCE) and diphenylamine (DPA). cDCE is a suspected carcinogen that frequently accumulates due to transformation of perchloroethene and trichloroethene at many contaminated sites. Polaromonas sp. strain JS666 is the only isolate able to use cDCE as the growth substrate, but the degradation mechanism was unknown. In this study, the degradation pathway of cDCE by strain JS666 and the genes involved were determined by using heterologous gene expression, inhibition studies, enzyme assays, and analysis of intermediates. The requirement of oxygen for cDCE degradation and inhibition of cDCE degradation by cytochrome P450 specific inhibitors suggested that cytochrome P450 monooxygenase catalyzes the initial steps of cDCE degradation. The finding was supported by the observation that an E. coli recombinant expressing cytochrome P450 monooxygenase catalyzes the transformation of cDCE to dichloroacetaldehyde and small amounts of the epoxide. Both the transient accumulation of dichloroacetaldehyde in cDCE degrading cultures and dichloroacetaldehyde dehydrogenase activities in cell extracts of JS666 further support a pathway involving the degradation of cDCE through dichloroacetaldehyde. Molecular phylogeny of the cytochrome P450 gene and organization of neighboring genes suggest that the cDCE degradation pathway evolved in a progenitor capable of degrading dichloroacetaldehyde by the recruitment of the cytochrome P450 monooxygenase gene from alkane assimilating bacteria. The discovery provides insight about the evolution of the aerobic cDCE biodegradation pathway and sets the stage for field applications.
DPA has been widely used as a precursor of dyes, pesticides, pharmaceuticals, and photographic chemicals and as a stabilizer for explosives, but little was known about the biodegradation of the compound. Therefore, bacteria able to use DPA as the growth substrate were isolated by selective enrichment from DPA-contaminated sediment and the degradation pathway and the genes that encode the enzymes were elucidated. Transposon mutagenesis, the sequence similarity of putative open reading frames to those of well characterized dioxygenases, and 18O2 experiments support the conclusion that the initial reaction in DPA degradation is catalyzed by a multi-component ring-hydroxylating dioxygenase. Aniline and catechol produced from the initial reaction of DPA degradation are then completely degraded via the common aniline degradation pathway. Molecular phylogeny and organization of the genes involved were investigated to provide insight about the evolution of DPA biodegradation.
The fate and transport of toxic chemicals are of a great concern at several historically contaminated sites where anoxic contaminant plumes emerge into water bodies. The release of toxic chemicals to overlying water poses a potential source of environmental exposure. Bench scale studies were conducted to evaluate the impact of biodegradation on the transport of toxic chemicals across the sediment/water interface. These studies demonstrated that substantial populations of bacteria associated with organic detritus at the interface rapidly biodegrade toxic chemicals as they migrate from contaminated sediment to overlying water, suggesting that the natural attenuation processes serve as a remedial strategy for contaminated sediments and protect the overlying water.
| Identifer | oai:union.ndltd.org:GATECH/oai:smartech.gatech.edu:1853/33974 |
| Date | 02 April 2010 |
| Creators | Shin, Kwanghee |
| Publisher | Georgia Institute of Technology |
| Source Sets | Georgia Tech Electronic Thesis and Dissertation Archive |
| Detected Language | English |
| Type | Dissertation |
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