title: Improving protein extraction and separation methods for investigating the metaproteome of anaerobic benzene communities within sediments.
abstract: BTEX compounds such as benzene are frequent soil and groundwater contaminants that are easily biodegraded under oxic conditions by bacteria. In contrast, benzene is rather recalcitrant under anaerobic conditions. The analysis of anoxic degradation is often hampered by difficult sampling conditions, limited amounts of biomass and interference of matrix compounds with proteomic approaches. In order to improve the procedure for protein extraction we established a scheme consisting of the following steps: dissociation of cells from lava granules, cell lysis by ultrasonication and purification of proteins by phenol extraction. The 2D-gels revealed a resolution of about 240 proteins spots and the spot patterns showed strong matrix dependence, but still differences were detectable between the metaproteomes obtained after growth on benzene and benzoate. Using direct data base search as well as de novo sequencing approaches we were able to identify several proteins. An enoyl-CoA hydratase with cross species homology to Azoarcus evansii, is known to be involved in the anoxic degradation of xenobiotics. Thereby the identification confirmed that this procedure has the capacity to analyse the metaproteome of an anoxic living microbial community.
authors.affiliation: Department of Proteomics, Helmholtz Centre for Environmental Research-UFZ, Permoserstr. 15, 04318 Leipzig, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Benndorf
authors.forename: Carsten
authors.initials: C
authors.lastname: Vogt
authors.forename: Nico
authors.initials: N
authors.lastname: Jehmlich
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Schmidt
authors.forename: Henrik
authors.initials: H
authors.lastname: Thomas
authors.forename: Gary
authors.initials: G
authors.lastname: Woffendin
authors.forename: Andrej
authors.initials: A
authors.lastname: Shevchenko
authors.forename: Hans-Hermann
authors.initials: HH
authors.lastname: Richnow
authors.forename: Martin
authors.initials: M
authors.lastname: von Bergen
mesh_terms: D000693:Anaerobiosis; D001419:Bacteria / Q000254:growth & development / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification*; D001554:Benzene / Q000378:metabolism*; D001565:Benzoates / Q000378:metabolism; D001673:Biodegradation, Environmental; D015180:Electrophoresis, Gel, Two-Dimensional; D019015:Geologic Sediments / Q000382:microbiology*; D013058:Mass Spectrometry; D056186:Metagenomics; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*
chemical_list: D001426:Bacterial Proteins; D001565:Benzoates; D020543:Proteome; D001554:Benzene