Characterization of biphenyl dioxygenase sequences and activities encoded by the metagenomes of highly polychlorobiphenyl-contaminated soils.

2.50
Hdl Handle:
http://hdl.handle.net/10033/245751
Title:
Characterization of biphenyl dioxygenase sequences and activities encoded by the metagenomes of highly polychlorobiphenyl-contaminated soils.
Authors:
Standfuss-Gabisch, Christine; Al-Halbouni, Djamila; Hofer, Bernd
Abstract:
Total extracted DNA from two heavily polychlorobiphenyl-contaminated soils was analyzed with respect to biphenyl dioxygenase sequences and activities. This was done by PCR amplification and cloning of a DNA segment encoding the active site of the enzyme. The translated sequences obtained fell into three similarity clusters (I to III). Sequence identities were high within but moderate or low between the clusters. Members of clusters I and II showed high sequence similarities with well-known biphenyl dioxygenases. Cluster III showed low (43%) sequence identity with a biphenyl dioxygenase from Rhodococcus jostii RHA1. Amplicons from the three clusters were used to reconstitute and express complete biphenyl dioxygenase operons. In most cases, the resulting hybrid dioxygenases were detected in cell extracts of the recombinant hosts. At least 83% of these enzymes were catalytically active. Several amino acid exchanges were identified that critically affected activity. Chlorobiphenyl turnover by the enzymes containing the prototype sequences of clusters I and II was characterized with 10 congeners that were major, minor, or not constituents of the contaminated soils. No direct correlations were observed between on-site concentrations and rates of productive dioxygenations of these chlorobiphenyls. The prototype enzymes displayed markedly different substrate and product ranges. The cluster II dioxygenase possessed a broader substrate spectrum toward the assayed congeners, whereas the cluster I enzyme was superior in the attack of ortho-chlorinated aromatic rings. These results demonstrate the feasibility of the applied approach to functionally characterize dioxygenase activities of soil metagenomes via amplification of incomplete genes.
Affiliation:
Division of Microbiology, Helmholtz-Zentrum für Infektionsforschung, Braunschweig, Germany.
Citation:
Characterization of biphenyl dioxygenase sequences and activities encoded by the metagenomes of highly polychlorobiphenyl-contaminated soils. 2012, 78 (8):2706-15 Appl. Environ. Microbiol.
Journal:
Applied and environmental microbiology
Issue Date:
Apr-2012
URI:
http://hdl.handle.net/10033/245751
DOI:
10.1128/AEM.07381-11
PubMed ID:
22327590
Type:
Article
Language:
en
ISSN:
1098-5336
Appears in Collections:
Publications of Dept. Chemical Biology (CBIO)

Full metadata record

DC FieldValue Language
dc.contributor.authorStandfuss-Gabisch, Christineen_GB
dc.contributor.authorAl-Halbouni, Djamilaen_GB
dc.contributor.authorHofer, Bernden_GB
dc.date.accessioned2012-09-25T08:16:37Z-
dc.date.available2012-09-25T08:16:37Z-
dc.date.issued2012-04-
dc.identifier.citationCharacterization of biphenyl dioxygenase sequences and activities encoded by the metagenomes of highly polychlorobiphenyl-contaminated soils. 2012, 78 (8):2706-15 Appl. Environ. Microbiol.en_GB
dc.identifier.issn1098-5336-
dc.identifier.pmid22327590-
dc.identifier.doi10.1128/AEM.07381-11-
dc.identifier.urihttp://hdl.handle.net/10033/245751-
dc.description.abstractTotal extracted DNA from two heavily polychlorobiphenyl-contaminated soils was analyzed with respect to biphenyl dioxygenase sequences and activities. This was done by PCR amplification and cloning of a DNA segment encoding the active site of the enzyme. The translated sequences obtained fell into three similarity clusters (I to III). Sequence identities were high within but moderate or low between the clusters. Members of clusters I and II showed high sequence similarities with well-known biphenyl dioxygenases. Cluster III showed low (43%) sequence identity with a biphenyl dioxygenase from Rhodococcus jostii RHA1. Amplicons from the three clusters were used to reconstitute and express complete biphenyl dioxygenase operons. In most cases, the resulting hybrid dioxygenases were detected in cell extracts of the recombinant hosts. At least 83% of these enzymes were catalytically active. Several amino acid exchanges were identified that critically affected activity. Chlorobiphenyl turnover by the enzymes containing the prototype sequences of clusters I and II was characterized with 10 congeners that were major, minor, or not constituents of the contaminated soils. No direct correlations were observed between on-site concentrations and rates of productive dioxygenations of these chlorobiphenyls. The prototype enzymes displayed markedly different substrate and product ranges. The cluster II dioxygenase possessed a broader substrate spectrum toward the assayed congeners, whereas the cluster I enzyme was superior in the attack of ortho-chlorinated aromatic rings. These results demonstrate the feasibility of the applied approach to functionally characterize dioxygenase activities of soil metagenomes via amplification of incomplete genes.en_GB
dc.language.isoenen
dc.rightsArchived with thanks to Applied and environmental microbiologyen_GB
dc.subject.meshCloning, Molecularen_GB
dc.subject.meshCluster Analysisen_GB
dc.subject.meshDioxygenasesen_GB
dc.subject.meshMetagenomeen_GB
dc.subject.meshMolecular Sequence Dataen_GB
dc.subject.meshPhylogenyen_GB
dc.subject.meshPolychlorinated Biphenylsen_GB
dc.subject.meshPolymerase Chain Reactionen_GB
dc.subject.meshSequence Analysis, DNAen_GB
dc.subject.meshSequence Homology, Amino Aciden_GB
dc.subject.meshSoil Microbiologyen_GB
dc.subject.meshSoil Pollutantsen_GB
dc.titleCharacterization of biphenyl dioxygenase sequences and activities encoded by the metagenomes of highly polychlorobiphenyl-contaminated soils.en
dc.typeArticleen
dc.contributor.departmentDivision of Microbiology, Helmholtz-Zentrum für Infektionsforschung, Braunschweig, Germany.en_GB
dc.identifier.journalApplied and environmental microbiologyen_GB
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