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Biodegradation of variable-chain-length n-alkanes in Rhodococcus opacus R7 and the involvement of an alkane hydroxylase system in the metabolism.

Zampolli J, Collina E, Lasagni M, Di Gennaro P - AMB Express (2014)

Bottom Line: This represents a difference in comparison with other Rhodococcus strains, in which a mixture of the two alcohols was observed.By GC-MSD analysis we also identified the monocarboxylic acid, confirming the terminal oxidation.Moreover, the alkB gene cluster from R. opacus R7 was isolated and its involvement in the n-alkane degradation system was investigated by the cloning of this genomic region into a shuttle-vector E. coli-Rhodococcus to evaluate the alkane hydroxylase activity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, Milano, 20126, Italy ; Department of Earth and Environmental Sciences, University of Milano-Bicocca, Milano, Italy.

ABSTRACT
Rhodococcus opacus R7 is a Gram-positive bacterium isolated from a polycyclic aromatic hydrocarbon contaminated soil for its versatile metabolism; indeed the strain is able to grow on naphthalene, o-xylene, and several long- and medium-chain n-alkanes. In this work we determined the degradation of n-alkanes in Rhodococcus opacus R7 in presence of n-dodecane (C12), n-hexadecane (C16), n-eicosane (C20), n-tetracosane (C24) and the metabolic pathway in presence of C12. The consumption rate of C12 was 88%, of C16 was 69%, of C20 was 51% and of C24 it was 78%. The decrement of the degradation rate seems to be correlated to the length of the aliphatic chain of these hydrocarbons. On the basis of the metabolic intermediates determined by the R7 growth on C12, our data indicated that R. opacus R7 metabolizes medium-chain n-alkanes by the primary alcohol formation. This represents a difference in comparison with other Rhodococcus strains, in which a mixture of the two alcohols was observed. By GC-MSD analysis we also identified the monocarboxylic acid, confirming the terminal oxidation. Moreover, the alkB gene cluster from R. opacus R7 was isolated and its involvement in the n-alkane degradation system was investigated by the cloning of this genomic region into a shuttle-vector E. coli-Rhodococcus to evaluate the alkane hydroxylase activity. Our results showed an increased biodegradation of C12 in the recombinant strain R. erythropolis AP (pTipQT1-alkR7) in comparison with the wild type strain R. erythropolis AP. These data supported the involvement of the alkB gene cluster in the n-alkane degradation in the R7 strain.

No MeSH data available.


Related in: MedlinePlus

Kinetic analyses ofn-dodecane (C12) (a),n-hexadecane (C16) (b),n-eicosane (C20) (c),n-tetracosane (C24) (d) degradation inR. opacusR7. Cells of R. opacus R7 were exposed to the n-alkane after growth on naphthalene and a flask each day was sacrified for the extraction and the GC-MSD determination of the residual hydrocarbon.
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Figure 1: Kinetic analyses ofn-dodecane (C12) (a),n-hexadecane (C16) (b),n-eicosane (C20) (c),n-tetracosane (C24) (d) degradation inR. opacusR7. Cells of R. opacus R7 were exposed to the n-alkane after growth on naphthalene and a flask each day was sacrified for the extraction and the GC-MSD determination of the residual hydrocarbon.

Mentions: To investigate the capacity of R. opacus R7 to degrade n-alkanes, kinetic runs were performed in M9 mineral medium supplemented with C12, C16, C20, C24 as the sole carbon and energy source; growth was followed at different times up to 72 h (Figure 1, a, b, c, d). The increase in biomass was quite the same for the four substrates, indeed OD600 reached about 0.5-0.7. GC-MSD analyses of n-alkane residues were performed after extraction from broths; the results showed that substrate consumption occurred parallel to biomass increase.


Biodegradation of variable-chain-length n-alkanes in Rhodococcus opacus R7 and the involvement of an alkane hydroxylase system in the metabolism.

Zampolli J, Collina E, Lasagni M, Di Gennaro P - AMB Express (2014)

Kinetic analyses ofn-dodecane (C12) (a),n-hexadecane (C16) (b),n-eicosane (C20) (c),n-tetracosane (C24) (d) degradation inR. opacusR7. Cells of R. opacus R7 were exposed to the n-alkane after growth on naphthalene and a flask each day was sacrified for the extraction and the GC-MSD determination of the residual hydrocarbon.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230829&req=5

Figure 1: Kinetic analyses ofn-dodecane (C12) (a),n-hexadecane (C16) (b),n-eicosane (C20) (c),n-tetracosane (C24) (d) degradation inR. opacusR7. Cells of R. opacus R7 were exposed to the n-alkane after growth on naphthalene and a flask each day was sacrified for the extraction and the GC-MSD determination of the residual hydrocarbon.
Mentions: To investigate the capacity of R. opacus R7 to degrade n-alkanes, kinetic runs were performed in M9 mineral medium supplemented with C12, C16, C20, C24 as the sole carbon and energy source; growth was followed at different times up to 72 h (Figure 1, a, b, c, d). The increase in biomass was quite the same for the four substrates, indeed OD600 reached about 0.5-0.7. GC-MSD analyses of n-alkane residues were performed after extraction from broths; the results showed that substrate consumption occurred parallel to biomass increase.

Bottom Line: This represents a difference in comparison with other Rhodococcus strains, in which a mixture of the two alcohols was observed.By GC-MSD analysis we also identified the monocarboxylic acid, confirming the terminal oxidation.Moreover, the alkB gene cluster from R. opacus R7 was isolated and its involvement in the n-alkane degradation system was investigated by the cloning of this genomic region into a shuttle-vector E. coli-Rhodococcus to evaluate the alkane hydroxylase activity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, Milano, 20126, Italy ; Department of Earth and Environmental Sciences, University of Milano-Bicocca, Milano, Italy.

ABSTRACT
Rhodococcus opacus R7 is a Gram-positive bacterium isolated from a polycyclic aromatic hydrocarbon contaminated soil for its versatile metabolism; indeed the strain is able to grow on naphthalene, o-xylene, and several long- and medium-chain n-alkanes. In this work we determined the degradation of n-alkanes in Rhodococcus opacus R7 in presence of n-dodecane (C12), n-hexadecane (C16), n-eicosane (C20), n-tetracosane (C24) and the metabolic pathway in presence of C12. The consumption rate of C12 was 88%, of C16 was 69%, of C20 was 51% and of C24 it was 78%. The decrement of the degradation rate seems to be correlated to the length of the aliphatic chain of these hydrocarbons. On the basis of the metabolic intermediates determined by the R7 growth on C12, our data indicated that R. opacus R7 metabolizes medium-chain n-alkanes by the primary alcohol formation. This represents a difference in comparison with other Rhodococcus strains, in which a mixture of the two alcohols was observed. By GC-MSD analysis we also identified the monocarboxylic acid, confirming the terminal oxidation. Moreover, the alkB gene cluster from R. opacus R7 was isolated and its involvement in the n-alkane degradation system was investigated by the cloning of this genomic region into a shuttle-vector E. coli-Rhodococcus to evaluate the alkane hydroxylase activity. Our results showed an increased biodegradation of C12 in the recombinant strain R. erythropolis AP (pTipQT1-alkR7) in comparison with the wild type strain R. erythropolis AP. These data supported the involvement of the alkB gene cluster in the n-alkane degradation in the R7 strain.

No MeSH data available.


Related in: MedlinePlus