The aim of the present research is to study the complex microbial adaptations induced by exposure to aromatic pollutants through a comprehensive proteomic analysis of A. radioresistens S13 including cytosolic, membrane, acidic and alkaline proteins and glycoproteins. Methods Cells were grown with either an aromatic (benzoate or phenol) or a non-aromatic (acetate) substrate as the sole carbon and energy source. Crude extracts were separated into cytosolic and membrane fractions by alkaline Na2CO3 treatment: 2DE was performed using 4-7 and 6-11 IPGstrips. Furthermore two methods for in-gel detection of glycoproteins were used. Spots were identified by N-terminal or de novo sequencing. Results Identified proteins belong to six functional classes: 1) Bioemulsification of hydrocarbons, including AlnA identified in both cytosolic and membrane extracts and present in two (a 39 kDa and a glycosylated 45 kDa forms) isoforms; 2) Outer (aspecific porin, BenP, PhoE) or inner (ABC transporters, Pho640, BenK) membrane transporters (some of these proteins being identified onto alkaline 2DE gels); 3) Defense mechanisms against aromatic toxicity (OHR, GroEL, TonB and ADH, all located in the cytoplasm); 4) Energy metabolism or pH homeostasis (ATP synthase, Na+/H+ antiporter); 5) Metabolism of polysaccharides and glycoproteins belonging to the capsule or the cell wall or extruded into the external environment (glycosyl transferases, SpsB); 6) Metabolism of lipids (ACP synthase). Conclusion Our results allow to outline a global picture of the bacterial response to aromatic compounds that includes modifications of the energetic state and of the cell surface hydrophobicity and the biosynthesis of stress proteins.
PHYSIOLOGICAL RESPONSES OF ACINETOBACTER RADIORESISTENS S13 TO AROMATIC MOLECULES DETECTED BY PROTEOMICS AND GLYCOPROTEOMICS
MAZZOLI, Roberto;BARELLO, CRISTINA;FATTORI, Paolo;GIUNTA, Carlo;PESSIONE, Enrica
2005-01-01
Abstract
The aim of the present research is to study the complex microbial adaptations induced by exposure to aromatic pollutants through a comprehensive proteomic analysis of A. radioresistens S13 including cytosolic, membrane, acidic and alkaline proteins and glycoproteins. Methods Cells were grown with either an aromatic (benzoate or phenol) or a non-aromatic (acetate) substrate as the sole carbon and energy source. Crude extracts were separated into cytosolic and membrane fractions by alkaline Na2CO3 treatment: 2DE was performed using 4-7 and 6-11 IPGstrips. Furthermore two methods for in-gel detection of glycoproteins were used. Spots were identified by N-terminal or de novo sequencing. Results Identified proteins belong to six functional classes: 1) Bioemulsification of hydrocarbons, including AlnA identified in both cytosolic and membrane extracts and present in two (a 39 kDa and a glycosylated 45 kDa forms) isoforms; 2) Outer (aspecific porin, BenP, PhoE) or inner (ABC transporters, Pho640, BenK) membrane transporters (some of these proteins being identified onto alkaline 2DE gels); 3) Defense mechanisms against aromatic toxicity (OHR, GroEL, TonB and ADH, all located in the cytoplasm); 4) Energy metabolism or pH homeostasis (ATP synthase, Na+/H+ antiporter); 5) Metabolism of polysaccharides and glycoproteins belonging to the capsule or the cell wall or extruded into the external environment (glycosyl transferases, SpsB); 6) Metabolism of lipids (ACP synthase). Conclusion Our results allow to outline a global picture of the bacterial response to aromatic compounds that includes modifications of the energetic state and of the cell surface hydrophobicity and the biosynthesis of stress proteins.
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