Switch to
| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
2
|
| pubmed:dateCreated |
1992-8-25
|
| pubmed:abstractText |
The rat and human recombinant soluble and membrane-bound catechol O-methyltransferase (S- and MB-COMT, respectively) were expressed using mammalian and baculovirus vectors. Low levels of rat and human S-COMT polypeptides were detected by immunoprecipitation in K-562 cell lines transfected with the S-COMT vectors. From K-562 cells transfected with the rat MB-COMT construct, both S- and MB-COMT recombinant proteins were detected by a rat COMT-specific anti-serum. Infection of lepidopteran Spodoptera frugiperda cells with recombinant S- or MB-COMT baculovirus constructs yielded high amounts of enzymically active and immunoreactive S- or MB-COMT proteins, respectively. Pulse/chase experiments with [35S]methionine-labelled insect cells infected with the MB-COMT baculovirus showed that the 30-kDa recombinant human MB-COMT polypeptide was not processed into the 25-kDa S-COMT form. Subcellular fractionations of insect cells, followed by immunoblotting with COMT antiserum, showed that recombinant S-COMT was found only in the soluble, cytoplasmic fraction, whereas MB-COMT resided both in soluble and membrane fractions. The recombinant MB-COMT sedimented in Percoll gradients at the density of 1.042 g/ml cosedimenting with the plasma-membrane marker. Fractionation and immunoblotting experiments on homogenized total rat brains indicated that the rat S-COMT (24 kDa) and some of the rat MB-COMT (28 kDa) was recovered in soluble fractions, whereas the microsomal material having COMT activity contained the MB-COMT polypeptide. The rat brain microsomal MB-COMT had a density of 1.042 g/ml in Percoll gradients, cosedimenting with the plasma-membrane and rough-endoplasmic-reticulum marker enzymes. The meta/para methylation ratio of dihydroxybenzoic-acid substrate by different recombinant and rat brain COMT-containing subcellular fractions was analysed.
|
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Jul
|
| pubmed:issn |
0014-2956
|
| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:day |
15
|
| pubmed:volume |
207
|
| pubmed:geneSymbol |
COMT
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
813-21
|
| pubmed:dateRevised |
2007-7-23
|
| pubmed:meshHeading |
pubmed-meshheading:1633830-Animals,
pubmed-meshheading:1633830-Baculoviridae,
pubmed-meshheading:1633830-Base Sequence,
pubmed-meshheading:1633830-Brain,
pubmed-meshheading:1633830-Catechol O-Methyltransferase,
pubmed-meshheading:1633830-Cell Compartmentation,
pubmed-meshheading:1633830-Cell Membrane,
pubmed-meshheading:1633830-Cloning, Molecular,
pubmed-meshheading:1633830-Humans,
pubmed-meshheading:1633830-Methylation,
pubmed-meshheading:1633830-Molecular Sequence Data,
pubmed-meshheading:1633830-Moths,
pubmed-meshheading:1633830-Oligodeoxyribonucleotides,
pubmed-meshheading:1633830-Polymerase Chain Reaction,
pubmed-meshheading:1633830-Precipitin Tests,
pubmed-meshheading:1633830-Rats,
pubmed-meshheading:1633830-Recombinant Proteins,
pubmed-meshheading:1633830-Solubility,
pubmed-meshheading:1633830-Tumor Cells, Cultured
|
| pubmed:year |
1992
|
| pubmed:articleTitle |
Expression of recombinant soluble and membrane-bound catechol O-methyltransferase in eukaryotic cells and identification of the respective enzymes in rat brain.
|
| pubmed:affiliation |
Orion-Farmos Pharmaceuticals, Orion Research Center, Helsinki, Finland.
|
| pubmed:publicationType |
Journal Article,
In Vitro
|