Degradation of Diphenyl Ether in Sphingobium phenoxybenzoativorans SC_3 Is Initiated by a Novel Ring Cleavage Dioxygenase.

Sphingobium phenoxybenzoativorans SC_3 degrades and utilizes diphenyl ether (DE) or 2-carboxy-DE as its sole carbon and energy source. In this study, we report the degradation of DE and 2-carboxy-DE initiated by a novel ring cleavage angular dioxygenase (diphenyl ether dioxygenase [Dpe]) in the strain. Dpe functions at the angular carbon and its adjacent carbon (C-1a, C-2) of a benzene ring in DE (or the 2-carboxybenzene ring in 2-carboxy-DE) and cleaves the C-1a-C-2 bond (decarboxylation occurs simultaneously for 2-carboxy-DE), yielding 2,4-hexadienal phenyl ester, which is subsequently hydrolyzed to muconic acid semialdehyde and phenol. Dpe is a type IV Rieske non-heme iron oxygenase (RHO) and consists of three components: a hetero-oligomer oxygenase, a [2Fe-2S]-type ferredoxin, and a glutathione reductase (GR)-type reductase. Genetic analyses revealed that dpeA1A2 plays an essential role in the degradation and utilization of DE and 2-carboxy-DE in S. phenoxybenzoativorans SC_3. Enzymatic study showed that transformation of 1 molecule of DE needs two molecules of oxygen and two molecules of NADH, supporting the assumption that the cleavage of DE catalyzed by Dpe is a continuous two-step dioxygenation process: DE is dioxygenated at C-1a and C-2 to form a hemiacetal-like intermediate, which is further deoxygenated, resulting in the cleavage of the C-1a-C-2 bond to form one molecule of 2,4-hexadienal phenyl ester and two molecules of H2O. This study extends our knowledge of the mode and mechanism of ring cleavage of aromatic compounds.IMPORTANCE Benzene ring cleavage, catalyzed by dioxygenase, is the key and speed-limiting step in the aerobic degradation of aromatic compounds. As previously reported, in the ring cleavage of DEs, the benzene ring needs to be first dihydroxylated at a lateral position and subsequently dehydrogenated and opened through extradiol cleavage. This process requires three enzymes (two dioxygenases and one dehydrogenase). In this study, we identified a novel angular dioxygenase (Dpe) in S. phenoxybenzoativorans SC_3. Under Dpe-mediated catalysis, the benzene ring of DE is dioxygenated at the angular position (C-1a, C-2), resulting in the cleavage of the C-1a-C-2 bond to generate a novel product, 2,4-hexadienal phenyl ester. This process needs only one angular dioxygenase, Dpe. Thus, the ring cleavage catalyzed by Dpe represents a novel mechanism of benzene ring cleavage.

[Selection of denitrifying phosphorus-removing bacteria and its characteristic].

Using YG culture medium, combined with the experiments of blue-and white-colored screening, Albert staining and detection of phosphorus removal capacity, seven phosphorus-removing bacterium were isolated from activated sludge collected from aerobic biochemical pool of wastewater treating factory. After nitrate reducing and anoxic culturing experiment, a strain with high capability of denitrifying and phosphorus removing was selected. With the study of its physiology and biochemistry and the analysis of its 16S rRNA gene, this strain was identified as Pseudomonas grimontii, and named of C18. The phosphorus removal rate of C18 was 94.1% under aerobic condition in 24 hours (The phosphorus concentration in supernatant was 38.7 mg/L to 2.28 mg/L). The phosphorus removal rate of C18 was 88.3% under anoxic condition in 24 hours (the phosphorus concentration in supernatant was 44.5 mg/L to 5.21 mg/L), and the denitrifying rate of C18 was 83.4% under anoxic condition in 24 hours (the nitrate concentration in supernatant was 184.2 mg/L to 30.6 mg/L). The optimal temperature of C18 denitrifying and phosphorus removing was 30 degrees C; The optimal pH of C18 denitrifying and phosphorus removing was 7.5.