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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
40
|
| pubmed:dateCreated |
1995-11-14
|
| pubmed:abstractText |
The CobU protein of Salmonella typhimurium was overexpressed and purified to approximately 94% homogeneity. N-terminal sequencing of purified CobU confirmed the first 22 amino acids. In vitro assays showed that CobU has kinase and guanylyltransferase activities which catalyze the synthesis of adenosyl-cobinamide-GDP from adenosyl-cobinamide, via an adenosyl-cobinamide-phosphate intermediate. We present evidence that the transfer of the guanylyl moiety of GTP to adenosyl-cobinamide-phosphate proceeds via an phosphoramidate-linked, enzyme-guanylyl intermediate. In the presence of oxygen, kinase and guanylyltransferase activities of CobU were lost. Treatment of inactive CobU with dithiothreitol restored approximately 20% of the kinase and guanylyltransferase activities, indicating the involvement of sulfhydryl groups in enzyme activity. The sulfhydryl modifying agents 5,5'-dithiobis(2-nitrobenzoic acid) and N-ethylmaleimide abolished both CobU activities. Native CobU protein was a dimer (approximately 40 kDa) that functioned optimally at pH 8.8-9.0 and 37 degrees C. Substrates and kinetic parameters for both activities were determined. The preferred corrinoid substrate for this enzyme was adenosyl-cobinamide. In vitro experiments are consistent with previous genetic studies which had suggested that adenosyl-cobinamide was the preferred substrate of CobU, and that CobU functioned more efficiently in the absence of oxygen.
|
| pubmed:grant | |
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/5'-Guanylic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Cobamides,
http://linkedlifedata.com/resource/pubmed/chemical/DNA, Bacterial,
http://linkedlifedata.com/resource/pubmed/chemical/Dithionitrobenzoic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Multienzyme Complexes,
http://linkedlifedata.com/resource/pubmed/chemical/Nucleotidyltransferases,
http://linkedlifedata.com/resource/pubmed/chemical/Pentosyltransferases,
http://linkedlifedata.com/resource/pubmed/chemical/nicotinate-nucleotide-dimethylbenzim...
|
| pubmed:status |
MEDLINE
|
| pubmed:month |
Oct
|
| pubmed:issn |
0021-9258
|
| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:day |
6
|
| pubmed:volume |
270
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
23560-9
|
| pubmed:dateRevised |
2007-11-14
|
| pubmed:meshHeading |
pubmed-meshheading:7559521-5'-Guanylic Acid,
pubmed-meshheading:7559521-Amino Acid Sequence,
pubmed-meshheading:7559521-Base Sequence,
pubmed-meshheading:7559521-Cobamides,
pubmed-meshheading:7559521-DNA, Bacterial,
pubmed-meshheading:7559521-Dithionitrobenzoic Acid,
pubmed-meshheading:7559521-Enzyme Stability,
pubmed-meshheading:7559521-Gene Expression,
pubmed-meshheading:7559521-Genes, Bacterial,
pubmed-meshheading:7559521-Hydrogen-Ion Concentration,
pubmed-meshheading:7559521-Kinetics,
pubmed-meshheading:7559521-Molecular Sequence Data,
pubmed-meshheading:7559521-Multienzyme Complexes,
pubmed-meshheading:7559521-Mutagenesis, Site-Directed,
pubmed-meshheading:7559521-Nucleotidyltransferases,
pubmed-meshheading:7559521-Pentosyltransferases,
pubmed-meshheading:7559521-Protein Conformation,
pubmed-meshheading:7559521-Salmonella typhimurium,
pubmed-meshheading:7559521-Substrate Specificity
|
| pubmed:year |
1995
|
| pubmed:articleTitle |
Purification and characterization of the bifunctional CobU enzyme of Salmonella typhimurium LT2. Evidence for a CobU-GMP intermediate.
|
| pubmed:affiliation |
Department of Bacteriology, University of Wisconsin, Madison 53706, USA.
|
| pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.,
Research Support, Non-U.S. Gov't
|