The restricted metabolism of the obligate organohalide respiring bacterium Dehalobacter restrictus: lessons from tiered functional genomics

Abstract

Dehalobacter restrictus strain PER-K23 is an obligate organohalide respiring bacterium, which displays extremely narrow metabolic capabilities. It grows only via coupling energy conservation to anaerobic respiration of tetra- and trichloroethene with hydrogen as sole electron donor. Dehalobacter restrictus represents the paradigmatic member of the genus Dehalobacter , which in recent years has turned out to be a major player in the bioremediation of an increasing number of organohalides, both in situ and in laboratory studies. The recent elucidation of the D. restrictus genome revealed a rather elaborate genome with predicted pathways that were not suspected from its restricted metabolism, such as a complete corrinoid biosynthetic pathway, the Wood–Ljungdahl (WL) pathway for CO 2 fixation, abundant transcriptional regulators and several types of hydrogenases. However, one important feature of the genome is the presence of 25 reductive dehalogenase genes, from which so far only one, pceA , has been characterized on genetic and biochemical levels. This study describes a multi-level functional genomics approach on D. restrictus across three different growth phases. A global proteomic analysis allowed consideration of general metabolic pathways relevant to organohalide respiration, whereas the dedicated genomic and transcriptomic analysis focused on the diversity, composition and expression of genes associated with reductive dehalogenases.

Genomic, Proteomic, and Biochemical Analysis of the Organohalide Respiratory Pathway in Desulfitobacterium dehalogenans

“…Triplicate samples were prepared for proteomic analysis as described before (38), with the modification that the protein samples were run over only a short distance, approximately 1 cm, in the SDS-PAGE gel prior to in-gel trypsin digestion of proteins. This was done to be able to cut and process entire lanes as single samples.…”

“…This was done to be able to cut and process entire lanes as single samples. Detection, identification, and relative quantification of proteins were done as described before (38). In brief, trypsin-digested peptides were run on a Proxeon LTQ-Orbitrap XL nano-liquid chromatography mass spectrometer (nLC-MS), and all spectral data obtained were analyzed with MaxQuant v.1.3.0.5 (39) using a protein database generated from the D. dehalogenans genome.…”

Overview of organohalide-respiring bacteria and a proposal for a classification system for reductive dehalogenases

“…The genome of Dehalogenimonas lykanthroporepellens strain BL-DC-9, the only sequenced Dehalogenimonas, contains 19 rdhA genes [17,66]. The presence of multiple rdhA genes is also a feature in the Dehalobacter genus, with up to 24 complete putative rdhA genes within sequenced Dehalobacter genomes [21,22,67].…”

Functional Genotyping of Sulfurospirillum spp. in Mixed Cultures Allowed the Identification of a New Tetrachloroethene Reductive Dehalogenase

“…The majority of currently known OHRB carry multiple rdhA genes, i.e., 1 to 36 genes, depending on the strain (12). Several studies have demonstrated that the presence of multiple nonidentical rdhA genes is a typical feature of OHRB (13)(14)(15). This suggests that the substrate range of the OHRB may be far greater than previously believed (16).…”