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0-Demethylation by the homoacetogenic anaerobe Holophaga foetida studied by a new photometric methylation assay using electrochemically produced cob(1)alamin

0-Demethylation by the homoacetogenic anaerobe Holophaga foetida studied by a new photometric methylation assay using electrochemically produced cob(1)alamin

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KREFT, Jan-Ulrich, Bernhard SCHINK, 1994. 0-Demethylation by the homoacetogenic anaerobe Holophaga foetida studied by a new photometric methylation assay using electrochemically produced cob(1)alamin. In: European Journal of Biochemistry. 226(3), pp. 945-951. ISSN 0014-2956. eISSN 1432-1033. Available under: doi: 10.1111/j.1432-1033.1994.00945.x

@article{Kreft19940Deme-7791, title={0-Demethylation by the homoacetogenic anaerobe Holophaga foetida studied by a new photometric methylation assay using electrochemically produced cob(1)alamin}, year={1994}, doi={10.1111/j.1432-1033.1994.00945.x}, number={3}, volume={226}, issn={0014-2956}, journal={European Journal of Biochemistry}, pages={945--951}, author={Kreft, Jan-Ulrich and Schink, Bernhard} }

<rdf:RDF xmlns:dcterms="http://purl.org/dc/terms/" xmlns:dc="http://purl.org/dc/elements/1.1/" xmlns:rdf="http://www.w3.org/1999/02/22-rdf-syntax-ns#" xmlns:bibo="http://purl.org/ontology/bibo/" xmlns:dspace="http://digital-repositories.org/ontologies/dspace/0.1.0#" xmlns:foaf="http://xmlns.com/foaf/0.1/" xmlns:void="http://rdfs.org/ns/void#" xmlns:xsd="http://www.w3.org/2001/XMLSchema#" > <rdf:Description rdf:about="https://kops.uni-konstanz.de/rdf/resource/123456789/7791"> <dc:rights>deposit-license</dc:rights> <foaf:homepage rdf:resource="http://localhost:8080/jspui"/> <dc:language>eng</dc:language> <dcterms:issued>1994</dcterms:issued> <dcterms:abstract xml:lang="eng">The previously studied complete methyl transfer sequence of tetrahydrofolate-dependent 0-demethylation catalyzed by Holophaga foetida strain TMBS4 extracts was separated into two steps using cobalamins as non-physiological substrates : electrochemically produced cob(1)alamin served as methyl acceptor for phenyl methyl ether demethylation, yielding methylcob(1II)alamin (reaction I), and methylcob(1II)alamin served as donor for tetrahydrofolate methylation, yielding 5-methyl tetrahydrofolate (reaction 11). Both reactions were measured with a new and direct photometric assay of cob(1)alamin methylation (or the reverse reaction) at 540 nm, the isosbestic wavelength of the cob(II)alamin/cob(I)alamin redox couple = 4.40 mM-' . cm-I). The rates of reactions I and I1 were proportional to protein concentration, unlike the complete reaction sequence. Small components of cell extract did not affect activity of reactions I and 11. Isovanillate demethylation by extracts of syringate-grown cells (reaction I) required reductive activation by cob(1)alamin and was inhibited and inactivated by cob(II)alamin, indicating that the reaction mechanism was a nucleo-philic attack of an enzyme-bound corrinoid in the reduced Co(1) state on the methyl carbon of the ether, rather than a radical attack. Only phenyl methyl ethers were demethylated; demethylation rates were enhanced by ortho-hydroxyl or para-carboxyl groups, but reduced by additional meta substituents. The rate of isovanillate demethylation was 81 nmol . min-' . (mg protein)-' [0.76 mM cob(I)alamin] and apparent kinetic constants for cob(1)alamin were: K, = 1.2 mM, V,,, = 220 nmol . min-' . (mg protein)-', and V,,,,,/K,,, = 180 nmol . min-' . (mg protein)-' . mN-'. 3,5-Dihydroxy-anisole demethylation by extracts of 3,5-dihydroxyanisole-grown cells (also reaction I) was much slower. Reaction I1 did not require activation; specific activity and the specificity constant for methylcob(II1)alamin were much lower.</dcterms:abstract> <dc:format>application/pdf</dc:format> <dcterms:rights rdf:resource="http://nbn-resolving.org/urn:nbn:de:bsz:352-20140905103416863-3868037-7"/> <dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/7791/1/1994_Kreft_945_951.pdf"/> <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/rdf/resource/123456789/28"/> <dc:contributor>Kreft, Jan-Ulrich</dc:contributor> <bibo:uri rdf:resource="http://kops.uni-konstanz.de/handle/123456789/7791"/> <dc:creator>Schink, Bernhard</dc:creator> <void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/> <dc:creator>Kreft, Jan-Ulrich</dc:creator> <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/rdf/resource/123456789/28"/> <dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/7791/1/1994_Kreft_945_951.pdf"/> <dc:contributor>Schink, Bernhard</dc:contributor> <dcterms:title>0-Demethylation by the homoacetogenic anaerobe Holophaga foetida studied by a new photometric methylation assay using electrochemically produced cob(1)alamin</dcterms:title> <dcterms:bibliographicCitation>First publ. in: European Journal of Biochemistry 226 (1994), 3, pp. 945-951</dcterms:bibliographicCitation> <dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2011-03-24T17:37:34Z</dcterms:available> <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2011-03-24T17:37:34Z</dc:date> </rdf:Description> </rdf:RDF>

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