Functional proteomic view of metabolic regulation in “Aromatoleum aromaticum” strain EbN1

Abstract

The denitrifying "Aromatoleum aromaticum" strain EbN1 utilizes a wide range of aromatic and nonaromatic compounds under anoxic and oxic conditions. The recently determined genome revealed corresponding degradation pathways and predicted a fine-tuned regulatory network. In this study, differential proteomics (2-D DIGE and MS) was used to define degradation pathway-specific subproteomes and to determine their growth condition dependent regulation. Differential protein profiles were determined for cultures adapted to growth under 22 different substrate and redox conditions. In total, 354 different proteins were identified, 199 of which displayed significantly changed abundances. These regulated proteins mainly represented enzymes of the different degradation pathways, and revealed different degrees of growth condition specific regulation. In case of three substrate conditions (e.g. phenylalanine, anoxic), proteins previously predicted to be involved in their degradation were apparently not involved (e.g. Pdh, phenylacetaldehyde dehydrogenase). Instead, previously not considered proteins were specifically increased in abundance (e.g. EbA5005, predicted aldehyde:ferredoxin oxidoreductase), shedding new light on the respective pathways. Moreover, strong evidence was obtained for thus far unpredicted degradation pathways of three hitherto unknown substrates (e.g. o-aminobenzoate, anoxic). Comparing all identified regulated and nonregulated proteins provided first insights into regulatory hierarchies of special degradation pathways versus general metabolism in strain EbN1.

Solvent Stress Response of the Denitrifying Bacterium “ Aromatoleum aromaticum ” Strain EbN1

“…In addition, strain EbN1 also utilizes p-cresol, phenol, and some other polar aromatic compounds under anoxic conditions. Recently, the complete genome sequence of strain EbN1 was determined, allowing detailed reconstruction of its metabolic network and in-depth proteomic analysis of substrate-dependent regulation of individual degradation pathways (28,35,54). Strain EbN1 represents a promising model organism for in situ processes, since toluene-amended microcosms from oil-contaminated and pristine sites displayed rapid consumption of nitrate (31) and contained high numbers of bacteria related to the phylogenetic cluster harboring strain EbN1 (20,31).…”

“…Cropped images were analyzed with the DeCyder software (version 5.0; GE Healthcare). The parameters used for codetection of spots were described recently (54). Spot matching was manually controlled for differentially regulated spots, which had to fulfill the following criteria: average ratio (fold change) of less than Ϫ2.5-fold or more than 2.5-fold, t test value of less than 10…”

Proteomics of marine bacteria

“…Varying levels of light intensity iTRAQ and LC-M/MS [43] Heterotrophy/photoautotrophy 2-D PAGE, MALDI-MS and LC-MS/MS [50] Nutrient availability/different C-and energy sources [68,74,75] Growth phase/cell cycle…”

“…A comprehensive proteome study of the Aromatoleum aromaticum strain EbN1 allowed for the exploration of the metabolic pathways, which are utilized by this denitrifying bacterium for the degradation of a wide range of aromatic and nonaromatic compounds under anoxic and oxic conditions [75]. The authors used 2-D DIGE analysis techniques to define degradation pathway-specific subproteomes and to determine their growth condition-dependent regulation.…”

“…In total, 354 different proteins could be identified, 199 of which displayed significantly changed abundances. The results of Wöhlbrand et al [75] revealed so far unpredicted degradation pathways of three hitherto unknown substrates.…”