Biochemical Characterization of Inducible ‘Reductase’ Component of Benzoate Dioxygenase and Phthalate Isomer Dioxygenases from Pseudomonas aeruginosa strain PP4

Abstract

The first step involved in the degradation of phthalate isomers (phthalate, isophthalate and terephthalate) is the double hydroxylation by respective aromatic-ring hydroxylating dioxygenases. These are two component enzymes consisting of 'oxygenase' and 'reductase' components. Soil isolate Pseudomonas aeruginosa strain PP4 degrades phthalate isomers via protocatechuate and benzoate via catechol 'ortho' ring cleavage pathway. Metabolic studies suggest that strain PP4 has carbon source-specific inducible phthalate isomer dioxygenase and benzoate dioxygenase. Thus, it was of interest to study the properties of reductase components of these enzymes. Reductase activity from phthalate isomer-grown cells was 3-5-folds higher than benzoate grown cells. In-gel activity staining profile showed a reductase activity band of R f 0.56 for phthalate isomer-grown cells as compared to R f 0.73 from benzoate-grown cells. Partially purified reductase components from phthalate isomer grown cells showed K m in the range of 30-40 μM and V max = 34-48 μmol min(-1) mg(-1). However, reductase from benzoate grown cells showed K m = 49 μM and V max = 10 μmol min(-1) mg(-1). Strikingly similar molecular and kinetic properties of reductase component from phthalate isomer-grown cells suggest that probably the same reductase component is employed in three phthalate isomer dioxygenases. However, reductase component is different, with respect to kinetic properties and zymogram analysis, from benzoate-grown cells when compared to that from phthalate isomer grown cells of PP4.

Catalytic and Spectroscopic Properties of the Halotolerant Soluble Methane Monooxygenase Reductase from Methylomonas methanica MC09

“…Interesting, the turnover rate was a magnitude higher than MmoC from M. capsulatus (Bath), [26] and three times higher than the related benzoate oxygenase reductase from Pseudomonas aeruginosa [27] with both dichlorophenylindophenol (DCIP) as the one electron acceptor. Considering the number of transferred electrons from the two‐electron donor NADH to the one‐electron acceptor benzyl viologen in case of MmoC and to the two‐electron acceptor DCIP, the MmoC activity is in the same range of benzoate oxygenase reductase, but still five times higher than MmoC from M. capsulatus (Bath), demonstrating that Mm MmoC is an attractive candidate for biotechnological approaches in combination with sMMO.…”

Biodegradation of phthalic acid esters by a newly isolated Mycobacterium sp. YC-RL4 and the bioprocess with environmental samples

“…Aerobic degradation of PAEs seems to be more efficient than is anaerobic process (Jiao et al 2013;Chen et al 2014). Isolated PAEs degrading bacteria mainly belong to the Pseudomonas, Gordonia, Arthrobacter, and Rhodococcus genera (Zeng et al 2008;Wu et al 2010;Yang et al 2013;Karandikar et al 2015). However, most of the reports focused on one special substrate and the degradation conditions, which does not meet the natural situation.…”

Microbial degradation of phthalates: biochemistry and environmental implications

“…The initial dihydroxylation of PA isomers has been studied most intensively with PA as substrate. Its dihydroxylation to cis ‐dihydrodiol‐PA is catalysed by two‐component Rieske non‐heme iron oxygenase systems comprising (i) a NAD(P)H‐dependent dioxygenase reductase component that harbours FMN and a plant‐type [2Fe‐2S] ferredoxin, and (ii) a non‐heme dioxygenase component binding a Rieske‐type [2Fe‐2S] cluster and an active site Fe atom [some selected references: (Batie et al ., ; Correll et al ., ; Tarasev and Ballou, ; Karandikar et al ., )]. There are two types of PA dioxygenases yielding either 4,5‐dihydro‐ cis ‐4,5‐dihydroxy‐PA (mostly in gram‐negative bacteria) or 3,4‐dihydro‐ cis ‐3,4‐dihydroxy‐PA (mostly in gram‐positive bacteria) (Fig.…”