Aminobacter sp. MSH1 Mineralizes the Groundwater Micropollutant 2,6-Dichlorobenzamide through a Unique Chlorobenzoate Catabolic Pathway

Abstract

2,6-dichlorobenzamide (BAM) is a major groundwater micropollutant posing problems for drinking water treatment plants (DWTPs) that depend on groundwater intake. Aminobacter sp. MSH1 uses BAM as a sole source of carbon, nitrogen and energy and is considered a prime biocatalyst for groundwater bioremediation in DWTPs. Its use in bioremediation requires knowledge on its BAM-catabolic pathway which is currently restricted to the amidase BbdA converting BAM into 2,6-dichlorobenzoic acid (2,6-DCBA) and the mono-oxygenase BbdD transforming 2,6-DCBA in 2,6-dichloro-3-hydroxybenzoic acid. Here, we show that the 2,6-DCBA catabolic pathway is unique and differs substantially from catabolism of other chlorobenzoates. BbdD catalyses a second hydroxylation forming 2,6-dichloro-3,5-dihydroxybenzoic acid. Subsequently, glutathione-dependent dehalogenases (BbdI and BbdE) catalyse the thiolytic removal of a first chlorine. The remaining chlorine is then removed hydrolytically by a dehalogenase of the / hydrolase superfamily (BbdC). BbdC is the first enzyme in that superfamily associated with dehalogenation of chlorinated aromatics and appears to represent a new subtype within the / hydrolase dehalogenases. The activity of BbdC yields a unique trihydroxylated aromatic intermediate for ring cleavage that is performed by an extradiol dioxygenase (BbdF) producing 2,4,6-trioxoheptanedioic acid which is likely converted to Krebs cycle intermediates by BbdG.

Bioaugmented Sand Filter Columns Provide Stable Removal of Pesticide Residue From Membrane Retentate

“…MSH1 can completely mineralize BAM without accumulation of any metabolites (Albers et al, 2014). The degradation pathway is shown in Raes et al (2019). Previous sand filter studies analyzing for confirmed and possible BAM metabolites; 2,6dichlorobenzoic acid (2,6-DCBA), ortho-chlorobenzamide, and ortho-chlorobenzoic acid, never detected these in the effluent of filters augmented with Aminobacter sp.…”

Toward Improved Bioremediation Strategies: Response of BAM-Degradation Activity to Concentration and Flow Changes in an Inoculated Bench-Scale Sediment Tank

“…15 The complete catabolic degradation pathway has recently been elucidated in detail. 20 The conversion of BAM to 2,6dichlorobenzoic acid ( of the overall process, 21 whereas further transformation of 2,6-DCBA is comparatively rapid in sand filters. 6 However, a specific challenge of using Aminobacter sp.…”

The complete genome of 2,6-dichlorobenzamide (BAM) degrader Aminobacter sp. MSH1 suggests a polyploid chromosome, phylogenetic reassignment, and functions of plasmids

“…Catabolic genes on pBAM1 and pBAM2 enable MSH1 to mineralize the groundwater micropollutant BAM and use it as a source of carbon, nitrogen, and energy for growth. The amidase BbdA on pBAM1 transforms BAM to 2,6-dichlorobenzoic acid (DCBA) 7 which is further metabolized by a series of catabolic enzymes encoded by pBAM2 8 , 9 . As previously discussed 8 , the gene bbdI encoding the gluthatione dependent thiolytic dehalogenase responsible for removal of one of the chlorines from BAM together with bbdJ encoding gluthatione reductase, occur on pBAM2 in three consecutive, perfect repeats followed by a fourth, imperfect repeat.…”

“…The first step of BAM-mineralization involves the hydrolysis of BAM to 2,6-dichlorobenzoic acid (2,6-DCBA) by the amidase BbdA encoded on the 41 kb IncP1-β plasmid pBAM1 7 . Further catabolism of 2,6-DCBA to central metabolism intermediates involves enzymes encoded on the 54 kb repABC family plasmid pBAM2 8 , 9 . The strain mineralizes BAM at trace concentrations 6 and invades biofilms of microbial communities of rapid sand filters used in drinking water treatment plants (DWTPs) 10 .…”