The isophthalate (IPA) degradation gene cluster (iphACBDR) responsible for the conversion of IPA into protocatechuate (PCA) was isolated from Comamonas sp. strain E6, which utilizes phthalate isomers as sole carbon and energy sources via the PCA 4,5-cleavage pathway. Based on amino acid sequence similarity, the iphA, iphC, iphB, iphD, and iphR genes were predicted to code for an oxygenase component of IPA dioxygenase (IPADO), a periplasmic IPA binding receptor, a 1,2-dihydroxy-3,5-cyclohexadiene-1,5-dicarboxylate (1,5-DCD) dehydrogenase, a reductase component of IPADO, and an IclR-type transcriptional regulator, respectively. The iphACBDR genes constitute a single transcriptional unit, and transcription of the iph catabolic operon was induced during growth of E6 on IPA. The iphA, iphD, and iphB genes were expressed in Escherichia coli. Crude IphA and IphD converted IPA in the presence of NADPH into a product which was transformed to PCA by IphB. These results suggested that IPADO is a two-component dioxygenase that consists of a terminal oxygenase component (IphA) and a reductase component (IphD) and that iphB encodes the 1,5-DCD dehydrogenase. Disruption of iphA and iphB resulted in complete loss of growth of E6 on IPA. Inactivation of iphD significantly affected growth on IPA, and the iphC mutant did not grow on IPA at neutral pH. These results indicated that the iphACBD genes are essential for the catabolism of IPA in E6. Disruption of iphR resulted in faster growth of E6 on IPA, suggesting that iphR encodes a repressor for the iph catabolic operon. Promoter analysis of the operon supported this notion.Phthalate isomers (o-phthalate [OPA], terephthalate [TPA], and isophthalate [IPA]) and their esters have been largely used as plasticizers. Moreover, they are considered potential starting compounds for the production of 2-pyrone-4,6-dicarboxylic acid (PDC), an intermediate metabolite in the protocatechuate (PCA) 4,5-cleavage pathway (28). PDC is a useful compound for synthesis of biodegradable and highly functional polymers, such as powerful adhesive agents (22,29,30).OPA degradation has been reported for many bacteria, including Burkholderia cepacia DBO1 (9), Mycobacterium vanbaalenii PYR-1 (43), Arthrobacter keyseri 12B (14), Terrabacter sp. DBF63 (19), Rhodococcus sp. DK17 (10), and Rhodococcus jostii RHA1 (32). Degradation of OPA is initiated by dihydroxylation of the aromatic ring by OPA 3,4-dioxygenase (10, 14), which is found mostly in Gram-positive bacteria, or by OPA 4,5-dioxygenase (9), which is found mostly in Gramnegative bacteria, which generates OPA dihydrodiols. These products are then transformed by rearomatization and decarboxylation by a dehydrogenase and a decarboxylase, respectively, yielding PCA. OPA dioxygenase from DBO1 has been purified (5). This enzyme is a multicomponent dioxygenase composed of a dioxygenase component and a reductase component. OPA dioxygenase requires Fe 2ϩ for activity and shows narrow substrate specificity with OPA. Microbial degradation of TPA has been characteriz...

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The tphA1 II and tphA2 II A3 II genes of Comamonas sp. E6 perhaps code for the terephthalate (TPA) 1,2-dioxygenase (TPADO). To characterize E6 TPADO, these genes were expressed in a His-tagged form in Escherichia coli, and the recombinant proteins were purified. TPADO activity was reconstituted from TphA1 II and TphA2 II A3 II, indicating that TPADO consists of a reductase (TphA1 II) and a terminal oxygenase component (TphA2 II and TphA3 II). TphA1(II) contains FAD, and the presence of a plant-type [2Fe-2S] cluster was suggested. These results indicate that TPADO is a class IB aromatic ring-hydroxylating dioxygenase. NADH and NADPH were effective as electron donors for TphA1 II, but NADPH appeared to be the physiological electron donor, based on the kinetic parameters. TPADO showed activity only toward TPA, and Fe2+ was required for it. The Km values for TPA and the Vmax were determined to be 72+/-6 microM and 9.87+/-0.06 U/mg respectively.

A real-time PCR quantification method for indigenous hydrocarbon-degrading bacteria (HDB) carrying the alkB gene in the soil environment was developed to investigate their distribution in soil. The detection limit of indigenous HDB by the method was 1 × 10(6) cells/g-soil. The indigenous HDB were widely distributed throughout the soil environment and ranged from 3.7 × 10(7) to 5.0 × 10(8) cells/g-soil, and the ratio to total bacteria was 0.1-4.3 %. The dynamics of total bacteria, indigenous HDB, and Rhodococcus erythropolis NDKK6 (carrying alkB R2) during bioremediation were analyzed. During bioremediation with an inorganic nutrient treatment, the numbers of these bacteria were slightly increased. The numbers of HDB (both indigenous bacteria and strain NDKK6) were gradually decreased from the middle stage of bioremediation. Meanwhile, the numbers of these bacteria were highly increased and were maintained during bioremediation with an organic nutrient. The organic treatment led to activation of not only the soil bacteria but also the HDB, so an efficient bioremediation was carried out.

It is important for conversational agents that manage multiparty conversations to recognize the group dynamics existing among the users. This paper proposes a method for estimating the conversational dominance of participants in group interactions. First, we conducted a Wizard-of-Oz experiment to collect conversational speech, and motion data. Then, we analyzed various paralinguistic speech and gaze behaviors to elucidate the factors that predict conversational dominance. Finally, by exploiting the speech and gaze data as estimation parameters, we created a regression model to estimate conversational dominance, and the multiple correlation coefficient of this model was 0.85.