Genetic and Structural Organization of the Aminophenol Catabolic Operon and Its Implication for Evolutionary Process

Abstract

The aminophenol (AP) catabolic operon in Pseudomonas putida HS12 mineralizing nitrobenzene was found to contain all the enzymes responsible for the conversion of AP to pyruvate and acetyl coenzyme A via extradiol meta cleavage of 2-aminophenol. The sequence and functional analyses of the corresponding genes of the operon revealed that the AP catabolic operon consists of one regulatory gene, nbzR, and the following nine structural genes, nbzJCaCbDGFEIH, which encode catabolic enzymes. The NbzR protein, which is divergently transcribed with respect to the structural genes, possesses a leucine zipper motif and a MarR homologous domain. It was also found that NbzR functions as a repressor for the AP catabolic operon through binding to the promoter region of the gene cluster in its dimeric form. A comparative study of the AP catabolic operon with other meta cleavage operons led us to suggest that the regulatory unit (nbzR) was derived from the MarR family and that the structural unit (nbzJCaCbDGFEIH) has evolved from the ancestral meta cleavage gene cluster. It is also proposed that these two functional units assembled through a modular type gene transfer and then have evolved divergently to acquire specialized substrate specificities (NbzCaCb and NbzD) and catalytic function (NbzE), resulting in the creation of the AP catabolic operon. The evolutionary process of the AP operon suggests how bacteria have efficiently acquired genetic diversity and expanded their metabolic capabilities by modular type gene transfer.Nitroaromatic compounds are widely used in the manufacture of dyes, drugs, explosives, and solvents, and massive amounts of their discharge have had detrimental effects on the environment. Much attention has been paid to the bioremediation of these compounds, and both aerobic and anaerobic degradation by microorganisms have been reported (13,16,45). As a typical recalcitrant nitroaromatic compound, nitrobenzene (NB) is known to be metabolized by aerobic bacteria through either an oxidative pathway (35) or a partial reductive pathway (20,22,34,38). In the partial reductive pathway, NB is converted into 2-aminophenol (AP), which is subsequently metabolized via a meta-like cleavage pathway. Several enzymes involved in NB catabolism of Pseudomonas pseudoalcaligenes JS45 were purified and characterized (21,23,24,31,44). However, despite intensive studies on the catabolic pathway of NB and characterization of the relevant enzymes, a detailed molecular basis of and a regulatory mechanism for NB catabolism remain yet to be elucidated. Recently, we reported the novel genetic organization of the NB catabolic gene clusters that are on the catabolic plasmids pNB1 and pNB2 in Pseudomonas putida HS12 (37). All the genes except for that of mutase, which was found on pNB2, were clustered on pNB1. Of the nbz (for nitrobenzene degradation) genes, the AP gene cluster was revealed to be a tightly regulated one.It has been generally accepted that horizontal gene transfer (HGT) has played an integral role in the dissemination of ...

TheclcElement ofPseudomonassp. Strain B13, a Genomic Island with Various Catabolic Properties

“…1D) (12), and AmnR, a repressor for the 2-AP pathway (in analogy to NbzR, a transcriptional repressor for the nbz genes of which the effector compound is unknown) (Fig. 1E) (45). In contrast to what we expected, we observed that the clc genes were induced in the presence of not only 3-CBA and 4-chlorobenzoate but also anthranilate, benzoate, and salicylate.…”

“…1E) consisted of a complete catabolic operon for 2-AP degradation via meta cleavage (31) that was similar to the amn gene cluster of Pseudomonas sp. strain AP-3 (61,62) and to parts of the nitrobenzene pathway genes of Pseudomonas fluorescens strain KU-7 (41) and P. putida HS12 (45). The amn operon on the clc element is formed by a putative transcriptional repressor (amnR, ORF22813), a ferredoxin-like protein (ORF23526), and eight amn genes, amnBACDFEHG ( Table 1. 1E).…”

Bacterial Transcriptional Regulators for Degradation Pathways of Aromatic Compounds

“…The nbz operon for aminophenol degradation on plasmid pNB1 in P. putida HS12 is regulated by NbzR, a MarR-type repressor protein (176). The chemical inducer for the pathway has not been identified, and it was proposed that another trans-acting factor might be involved in regulating pathway expression.…”

“…Instead, the protein possesses a putative leucine zipper motif (which is absent in other family members), which is thought to be responsible for the NbzR dimerization and DNA binding. Hence, Park and Kim suggested that the poor HTH motif of NbzR perhaps does not contribute to DNA binding (176). MarR-type regulators controlling catabolic operons are relatively small proteins.…”

“…Additional pathway control is exerted by the AadR regulator, a FNR-type regulator, under anaerobic conditions (55). The regulatory genes nbzR, cbaR, and hpcR are located upstream of and are transcribed in the opposite direction to their target operons (176,194,197). badR is located upstream of and transcribed in the same direction as its target operons but is separated from them by one additional gene (55).…”