Anaerobic Degradation of p -Ethylphenol by “ Aromatoleum aromaticum ” Strain EbN1: Pathway, Regulation, and Involved Proteins

Abstract

ABSTRACT The denitrifying “ Aromatoleum aromaticum ” strain EbN1 was demonstrated to utilize p -ethylphenol under anoxic conditions and was suggested to employ a degradation pathway which is reminiscent of known anaerobic ethylbenzene degradation in the same bacterium: initial hydroxylation of p -ethylphenol to 1-(4-hydroxyphenyl)-ethanol followed by dehydrogenation to p -hydroxyacetophenone. Possibly, subsequent carboxylation and thiolytic cleavage yield p -hydroxybenzoyl-coenzyme A (CoA), which is channeled into the central benzoyl-CoA pathway. Substrate-specific formation of three of the four proposed intermediates was confirmed by gas chromatographic-mass spectrometric analysis and also by applying deuterated p -ethylphenol. Proteins suggested to be involved in this degradation pathway are encoded in a single large operon-like structure (∼15 kb). Among them are a p -cresol methylhydroxylase-like protein (PchCF), two predicted alcohol dehydrogenases (ChnA and EbA309), a biotin-dependent carboxylase (XccABC), and a thiolase (TioL). Proteomic analysis (two-dimensional difference gel electrophoresis) revealed their specific and coordinated upregulation in cells adapted to anaerobic growth with p -ethylphenol and p -hydroxyacetophenone (e.g., PchF up to 29-fold). Coregulated proteins of currently unknown function (e.g., EbA329) are possibly involved in p -ethylphenol- and p -hydroxyacetophenone-specific solvent stress responses and related to other aromatic solvent-induced proteins of strain EbN1.