TOL plasmid pWW15 contains two nonhomologous, independently regulated catechol 2,3-oxygenase genes

Abstract

Pseudomonas putida MT15 contains a 250-kilobase-pair (kbp) TOL plasmid pWW15, encoding toluene and xylene catabolism, which undergoes large spontaneous deletions to give two classes of mutants with altered catabolic phenotypes (H. Keil and P. A. Williams, J. Gen. Microbiol, 131:1023-1033, 1985). Two structural genes for catechol 2,3-oxygenase (C230) were cloned from pWW15. The gene for C2301 was located on the 2.1-kbp XhoI fragment Xh, whereas that for C2301I was found on the 11.5-kbp BamHI fragment BJ. The two restriction fragments and the subcloned regions of them showed no similarity in the pattern of restriction digestion, nor did they hybridize with each other. The substrate specificities of the two enzymes were also substantially different. The two structural genes were separated on pWW15 by about 100 kbp. In plasmid pWW15-510 of a B5 mutant, the 90-kbp deletion in the plasmid removed most of the intervening DNA, but it also deleted 80% of the gene for C2301 from its 3' end. Thus, only C230II was expressed in the host MT15-510. Conversely, in RP4::pWW15 cointegrate plasmid pWW15-1003, only the C2301 gene was present. The expression of C230 activity from these two derivative plasmids and from the wild-type pWW15 showed that only C230I was induced by growth in the presence of m-toluate, whereas both activities were induced in the presence of m-xylene. These findings cast doubt on the earlier hypothesis that the deletions in B3 and B5 mutants remove a regulatory gene by which m-toluate induces the enzymes necessary for its own catabolism.

Metagenomics reveals diversity and abundance of meta‐cleavage pathways in microbial communities from soil highly contaminated with jet fuel under air‐sparging bioremediation

“…1A). The highest similarity (79-81% of identity) was observed with CdoII extradiol dioxygenase of P. putida MT15 (Keil et al, 1985). A subgroup of 11 gene fragments ( Fig.…”

Three different 2,3-dihydroxybiphenyl-1,2-dioxygenase genes in the gram-positive polychlorobiphenyl-degrading bacterium Rhodococcus globerulus P6

“…There are several examples of duplications of genes involved in biodegradation of toxic compounds, in particular of genes encoding ring cleavage enzymes. Nonhomologous catechol-2,3-dioxygenase genes have been found in A. eutrophus JMP222 (29,30), in Rhodococcus rhodochrous CTM (44), and in the TOL plasmid pWW15 of P. putida (32). Many naturally occurring TOL plasmids carry duplications of the upper-and/or lower-pathway operons, and/or the xylS and xylR regulatory genes (49); any selective advantage that these duplications might confer upon the plasmid has not yet been defined.…”

Homologies between plasmid and chromosomally-encoded benzoate-oxidizing genes in Pseudomonas putida

“…Previous studies on the meta-cleavage pathway operon of TOL plasmid pWWO (Harayama et al 1986; Harayama & Rekik 1990) have identified three polypeptides, termed XylXYZ, which together comprise the multi-enzyme complex for the toluate 1,2-dioxygenase. Both TOL plasmids pWWO and pWW53 show a highly conserved restriction pattern within the x y l X Y Z region (Keil et al 1985a) suggesting an identical location of the corresponding catabolic genes on the plasmids (Fig. 1).…”

“…putida P a w l . Recombinant plasmids used were pWW53-3403, pWW53-3555, pWW53-3606 (Keil et al 1985a) and pWW53-3825 (Keil et al 1987).…”