Enzymes involved in the anaerobic degradation of phenol by the sulfate-reducing bacterium Desulfatiglans anilini

Xiaoman Xie,Nicolai Müller

doi:10.1186/s12866-018-1238-0

Abstract

BackgroundThe sulfate-reducing bacterium Desulfatiglans anilini can grow with phenol as sole source of carbon and energy under strictly anaerobic, sulfate-reducing conditions. In the nitrate-reducing bacterium Thauera aromatica, the enzymes involved in phenol degradation have been well elucidated, whereas the anaerobic phenol degradation pathway by D. anilini was not studied in detail yet.ResultsThe pathway of anaerobic phenol degradation by the sulfate-reducing bacterium Desulfatiglans anilini was studied by identification of genes coding for phenylphosphate synthase (encoded by pps genes) and phenylphosphate carboxylase (encoded by ppc genes) in the genome of D. anilini, by analysis of the transcription and translation of pps-ppc genes, and by measurement of phenylphosphate synthase activity in cell-free extracts of phenol-grown cells. The majority of genes involved in phenol degradation were found to be organized in one gene cluster. The gene cluster contained genes ppsα (phenylphosphate synthase alpha subunit), ppsβ (phenylphosphate synthase beta subunit), ppcβ (phenylphosphate carboxylase beta subunit), as well as 4-hydroxybenzoyl-CoA ligase and 4-hydroxylbenzoyl-CoA reductase-encoding genes. The genes ppsγ (phenylphosphate synthase gamma subunit), ppcα (phenylphosphate carboxylase alpha subunit) and ppcδ (phenylphosphate carboxylase delta subunit) were located elsewhere in the genome of D. anilini, and no obvious homologue of ppcγ (phenylphosphate carboxylase gamma subunit) was found in the genome. Induction of genes pps and ppc during growth on phenol was confirmed by reverse transcription polymerase chain reaction. Total proteome analysis revealed that the abundance of enzymes encoded by the gene cluster under study was much higher in phenol-grown cells than that in benzoate-grown cells. In in-vitro enzyme assays with cell-free extracts of phenol-grown cells, phenylphosphate was formed from phenol in the presence of ATP, Mg2+, Mn2+, K+ as co-factors.ConclusionsThe genes coding for enzymes involved in the anaerobic phenol degradation pathway were identified in the sulfate-reducing bacterium D. anilini. The results indicate that the first steps of anaerobic phenol degradation in D. anilini are phosphorylation of phenol to phenylphosphate by phenylphosphate synthase and carboxylation of phenylphosphate by phenylphosphate carboxylase.

Highlights

The sulfate-reducing bacterium Desulfatiglans anilini can grow with phenol as sole source of carbon and energy under strictly anaerobic, sulfate-reducing conditions
We identified the catabolic enzymes and their genes involved in anaerobic degradation of phenol in the sulfate reducing bacterium Desulfatiglans anilini
Anaerobic growth with phenol or benzoate The growth of Desulfatiglans anilini on phenol or benzoate was investigated. 2 mM phenol or 2 mM benzoate was supplied to D. anilini cultures as the only source of electrons with Na2SO4 as the electron acceptor (Fig. 2)

Summary

Introduction

The sulfate-reducing bacterium Desulfatiglans anilini can grow with phenol as sole source of carbon and energy under strictly anaerobic, sulfate-reducing conditions. In the nitrate-reducing bacterium Thauera aromatica, the enzymes involved in phenol degradation have been well elucidated, whereas the anaerobic phenol degradation pathway by D. anilini was not studied in detail yet. Anaerobic phenol degradation has been studied in detail with the denitrifying bacterium Thauera aromatica. The enzyme involved in the first reaction is phenylphosphate synthase (encoded by pps genes), which converts phenol and ATP to phenylphosphate, AMP, and phosphate [5]. The α-subunit (70 kDa) containing a conserved histidine residue alone can catalyze the exchange of free [14C] phenol and the phenol moiety of phenylphosphate, but not the phosphorylation of phenol. The β-subunit (40 kDa) is required in the phosphorylation of phenol, which can transfer a diphosphoryl group to the conserved histidine residue in the α-subunit [6]. The reaction is stimulated by the addition of γ-subunit (24 kDa), but the exact function of the γ-subunit is unknown [5]

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