Naphthalene degradation plasmid

The aromatic degradative pathways receiving the most attention include those for toluene, benzoate, PCBs, and naphthalene. Five completely independent routes of toluene catabolism have been well characterized for aerobic bacteria (Figure 11.1). To date, all five pathways have been described in Pseudomonas and Burkholderia, despite the fact that at least two have been shown to be encoded by broad-host-range degradative plasmids. 

The reactions, described so far, are derived from tiie most extensively studied catabolic pathway for naphthalene, which is encoded by the NAH7 plasmid of Pseudomonas putida. But, the same sequence of reactions seems to take place in all naphthalene-degrading bacteria, which means that salicylate is a common intmmiediate in naphthalene catabolism. With respect to the enzymes involved in the breakdown of naphthalene, there seem to exist differences between the bacterial species, that can grow on naphtiialene-containing media. A recently isolated and described Rhodococcus sp., for example, seems to have a 1,2-dihydroxynaphthalene oxygenase that requires NADH [38]. 

Dunn NW, HM Dunn, RA Austen (1980) Evidence for the existence of two catabolic plasmids coding for the degradation of naphthalene. J Gen Microbiol 117 529-533. 

Carbazole, A-methylcarbazole, IV-ethylcarbazole, dibenzofuran, dibenzothiophene, fluorene, dibenzo-p-dioxin, phenoxathiin, phenoxazine, phenothiazine, xanthene, biphenyl, naphthalene, phenanthrene, anthracene, and fluoranthene could be transformed by E. coli, [314] which was transformed using a plasmid bearing the carAa, Ac, and Ad genes, and expressing only the carA-encoded proteins. Further work is needed to develop a final biocatalyst and to prove the advantages that this degradative pathway would incorporate in a refining bioprocess. 

That the traits of B. m aterium can be transferred to B. subtilis by plasmid transfer techniques has been established. These traits are increased resistance to naphthalene, parachlorbiphenyl, and dibenzofuran, and increased ability to degrade C-naphthalene. The transformants increased degradative abilities were demonstrated by the accumulation of greater quantitites of water soluble metabolite and the presence of a unique solvent soluble metabolite. 

While we did not test sufficient numbers of regenerated protoplasts to accurately determine the transformation frequency to aromatic hydrocarbon resistance, these frequencies were certainly high. Chang and Cohen (37) reported transformation frequencies of 10-80% using the same lysozyme-PEG method and similar quantities of plasmid DNA. The naphthalene transformants retained their degradative ability through at least six successive transfers. 

The plasmids and operons described above represent the most studied ones, but probably constitute a small fraction of the catabolic operons in bacteria. In one study, 43 bacterial strains (mostly Pseudomonas spp.) from different sources, shown to possess the ability to degrade aromatic and PAHs, were hybridized with probes of NAH and TOL plasmids as well as with genomic DNA of bacteria known to degrade a wide variety of PAHs. Only 14 strains that mineralized naphthalene and phenanthrene showed homology to one of the probes. The remaining isolates mineralized and/or oxidized various PAHs and hybridized with neither pure plasmids nor genomic DNA (Foght Westlake, 1991). 

The NAH and TOL plasmids which encode naphthalene and toluene degradation represent discrete, mobile genetic systems that are easily transferred and selected. This tendency to focus on systems according to the ease of genetic manipulation led... 

Figure 2. A) Genetic organization of the pKAl catabolic plasmid. Genes of the upper naphthalene regulatory system encode for proteins that mediate the conversion of naphthalene to salicylate. Salicylate is then further degraded to TCA cycle intermediates. B) In Pseudomonas fluorescens HK44, genes within the lower pathway were replaced with genes of the lux cassette to produce a bioluminescent bioreporter sensitive to naphthalene and salicylate.

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