Isolation and characterization of Achromobacter xylosoxidans T7 capable of degrading toluidine isomers.

A bacterial strain capable of utilizing toluidine isomers as its sole source of carbon and energy for growth was isolated from contaminated soil. The isolate was identified as Achromobacter xylosoxidans and was designated strain T7. Strain T7 differs from other toluidine-degrading strains with respect to the use of all three toluidine isomers even as an equimolar mixture. Additionally, strain T7 harbours the ability to use aniline, phenol, and cresols as growth substrates. Utilization of the toluidine isomers was demonstrated by an increase in the bacterial biomass concomitant with a decrease of the respective toluidine concentration in liquid medium with this compound as sole source of carbon and energy. No accumulation of any intermediate was detectable by HPLC-analyses. Results of oxygen uptake experiments with resting cells of strain T7 pre-grown on the respective toluidine and enzymatic investigations in cell-free extracts indicate the metabolization of the toluidines via the respective methylcatechols as intermediates. These compounds are substrates for the meta-cleavage pathway initiated by inducible catechol 2,3-dioxygenase found in toluidine-grown cells of strain T7.

Complex protein composition of isolated focal adhesions: a two-dimensional gel and database analysis.

Current ideas about the composition of the focal adhesion complexes in cultured cells are based mainly on indirect immunocytochemical data. We here report a two-dimensional (2-D) gel electrophoresis analysis of the focal adhesion associated-structures that remain in the growth substrate after removal of cells by mechanical shearing. Many proteins additional to the known adhesion proteins, and in higher abundance, could be identified. Using selective extraction procedures, employing detergent or gelsolin, these could be classified as either membrane-associated, actin-associated or both. Cross correlation of these polypeptode patterns with a 2-D gel database allowed identification of some proteins, not previously considered as resident of focal adhesions. The data point to a more complex make up of focal adhesions than formerly supposed.

Pseudomonas acidovorans: a bacterium capable of mineralizing 2-chloroaniline.

Prolonged adaptation of Ca-alginate immobilized cells of Pseudomonas acidovorans CA28 to a mixture of 3-chloroaniline (3-CA)1) and 2-CA and subsequently to 2-CA as sole substrate led to the isolation of another strain, termed CA50 with the additional capability of utilizing 2-CA as sole source of carbon, nitrogen, and energy. Batch-degradation of 190 mg/l of 2-CA at pH 6.1 by this newly isolated strain was achieved within 3 days, at higher concentrations up to 0.6 g/l increasing lag-phases and degradation periods were observed. Except chloride and ammonium no further metabolites were detectable in the medium. Mineralization of 2-CA proceeds via the modified ortho-cleavage pathway as demonstrated by the presence of catechol 1,2-dioxygenase (C120) activity, which is characterized by its substrate specificity and elution behaviour on DEAE-cellulose.

Characterization of isofunctional ring-cleaving enzymes in aniline and 3-chloroaniline degradation by Pseudomonas acidovorans CA28.

During degradation of aniline and 3-chloroaniline, respectively, by Pseudomonas acidovorans CA28, selective induction of two catechol 1,2-dioxygenases (C12O) was observed. C12O I activity was the sole ring-cleaving enzyme detectable in cell-free extracts after growth on aniline, while C12O II was exclusively found after growth on 3-chloroaniline. Both enzymes were clearly differentiated by their elution behaviour on DEAE-cellulose and their substrate specificities. For C12O I high activity was demonstrable only with unsubstituted catechol, while C12O II showed preference for and high affinity towards chlorinated catechols. Therefore, evidence of different ortho-cleavage enzymes in Pseudomonas acidovorans CA28 involved in aniline and 3-chloroaniline metabolism, respectively, is indicated.

Degradation of phenol and phenolic compounds by Pseudomonas putida EKII.

The phenol-degrading strain Pseudomonas putida EKII was isolated from a soil enrichment culture and utilized phenol up to 10.6 mM (1.0 g.l-1) as the sole source of carbon and energy. Furthermore, cresols, chlorophenols, 3,4-dimethylphenol, and 4-chloro-m-cresol were metabolized as sole substrates by phenol-grown resting cells of strain EKII. Under conditions of cell growth, degradation of these xenobiotics was achieved only in co-metabolism with phenol. Phenol hydroxylase activity was detectable in whole cells but not in cell-free extracts. The specificity of the hydroxylating enzyme was found during transformation of cresols and chlorophenols: ortho- and meta-substituted phenols were degraded via 3-substituted catechols, while degradation of para-substituted phenols proceeded via 4-substituted catechols. In cell-free extracts of phenol-grown cells a high level of catechol 2,3-dioxygenase as well as smaller amounts of 2-hydroxymuconic semialdehyde hydrolyase and catechol 1,2-dioxygenase were detected. The ring-cleaving enzymes were characterized after partial purification by DEAE-cellulose chromatography.

Isolation and characterization of a 2-(2,4-dichlorophenoxy) propionic acid-degrading soil bacterium.

The 2-(2,4-dichlorphenoxy)propionic acid (2,4-DP)-degrading bacterial strain MH was isolated after numerous subcultivations of a mixed culture obtained by soil-column enrichment and finally identified as Flavobacterium sp. Growth of this strain was supported by 2,4-DP (maximum specific growth rate 0.2 h-1) as well as by 2,4-dichlorophenoxyacetic acid (2,4-D), 4-(2,4-dichlorophenoxy)butyric acid (2,4-DB), and 2-(4-chloro-2-methylphenoxy)propionic acid (MCPP) as sole sources of carbon and energy under aerobic conditions. 2,4-DP-Grown cells (10(8] of strain MH degraded 2,4-dichlorophenoxyalkanoic acids, 2,4-dichlorophenol (2,4-DCP), and 4-chlorophenol at rates in the range of 30 nmol/h. Preliminary investigations indicate that cleavage of 2,4-DP results in 2,4-DCP, which is further mineralized via ortho-hydroxylation and ortho-cleavage of the resulting 3,5-dichlorocatechol.