DNA sequences of genes encoding Acinetobacter calcoaceticus protocatechuate 3,4-dioxygenase: evidence indicating shuffling of genes and of DNA sequences within genes during their evolutionary divergence

Abstract

The DNA sequence of a 2,391-base-pair HindIII restriction fragment of Acinetobacter calcoaceticus DNA containing the pcaCHG genes is reported. The Protocatechuate 3,4-dioxygenase ( Fig. 1; EC 1.13.11.3) plays an essential role in the utilization of numerous aromatic and hydroaromatic compounds via the P-ketoadipate pathway (45). Ubiquity of the pathway in the natural environment is indicated by the fact that the enzyme is a trait universally shared by fluorescent Pseudomonas species (46) and by members of the family Rhizobiaceae (39). The available evidence points to a single evolutionary origin for protocatechuate 3,4-dioxygenases formed by diverse bacteria (14). Other bacterial intradiol dioxygenases (Fig. 1) act upon catechol. Catechol oxygenase I (40) exhibits narrow substrate specificity and a relatively high Kcat (33) when compared with catechol oxygenase II (10), an enzyme that more readily accommodates chlorocatechol as a substrate and exhibits a relatively low Kcat (Fig. 1).The activity of intradiol dioxygenases depends on ferric ion which is ligated by two histidyl and two tyrosyl side chains within the catalytic subunit of the enzymes (41). Determination of the crystal structure of a Pseudomonas protocatechuate 3,4-dioxygenase has established the position of the iron-ligating histidyl and tyrosyl residues within the primary sequence of the a subunit (34). Like other protocatechuate 3,4-dioxygenases, the Pseudomonas enzyme contains in equimolar amounts the i subunit and an a subunit that contributes to substrate binding. Amino acid sequence comparisons (21,24)

Sequence analysis of the Pseudomonas sp. strain P51 tcb gene cluster, which encodes metabolism of chlorinated catechols: evidence for specialization of catechol 1,2-dioxygenases for chlorinated substrates

“…In contrast, the TfdC enzyme shows a higher rate of conversion with 3,5-DCC as a substrate, which is in agreement with the findings of Pieper et al (22). In A. eutrophus JMP134, 3,5-DCC occurs as an intermediate in the conversion of 2,4-dichlorophenoxyacetic acid by enzymes of the tfd pathway (5), indicating that this pathway is specialized for conversion of 3,5-DCC (22 (10,13,17). The data presented here reinforce the hypothesis that the intradiol dioxygenases have a common ancestor.…”

Cloning, sequencing, and expression of the Pseudomonas putida protocatechuate 3,4-dioxygenase genes

“…Although the genes for 3,4-PCD have been cloned and expressed from both P. cepacia (61,62) and Acinetobacter calcoaceticus (24), their use in mutational studies would be limited by the amount and type of physical and mechanistic data available for the 3,4-PCDs encoded by these genes. In addition, the recombinant enzymes from these sources were not overexpressed or extensively characterized.…”

Potential DNA slippage structures acquired during evolutionary divergence of Acinetobacter calcoaceticus chromosomal benABC and Pseudomonas putida TOL pWW0 plasmid xylXYZ, genes encoding benzoate dioxygenases

“…In contrast, genes for such isofunctional enzymes have diverged substantially, as indicated by differences in G+C content of about 15% (9,20,27). Such differences in G+C content may be attributed in part to directional pressure exerted by mutations within divergent cell lines (32).…”

“…The interaction of slipped DNA strands during mutation has been documented by demonstration that sequence-directed mutations create both deletions (3)(4)(5)(6) and repetitions of DNA sequence. Comparison of DNA sequences for oxygenative enzymes from A. calcoaceticus and P. putida suggested that slippage structures formed between misaligned DNA strands were formed by mutation and are maintained by mismatch repair during evolution (9,21).…”

“…Isofunctional enzymes for dissimilation of aromatic compounds in A. calcoaceticus and P. putida generally exhibit close evolutionary ancestry reflected in identities of amino acid sequence close to or exceeding 50% (9,23,38,39,41). In contrast, genes for such isofunctional enzymes have diverged substantially, as indicated by differences in G+C content of about 15% (9,20,27).…”