Linked Life Data

2549028

Source:http://linkedlifedata.com/resource/pubmed/id/2549028

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
25
pubmed:dateCreated
1989-10-11
pubmed:abstractText
The tumor necrosis factor (TNF) receptor of HeLa cells was solubilized in Triton X-100 and characterized by gel filtration, affinity labeling, and ligand blotting studies. Receptors solubilized with Triton X-100 eluted in gel filtration as a major peak of Mr = 330,000 and retained high affinity binding (KD = 0.25 nM). Affinity labeling of soluble receptor/125I-TNF complexes using the reversible, bifunctional bis[2-(succinimidooxycarbonyl-oxy)ethyl] sulfone resulted in the formation of cross-linked species of Mr = 310,000, 150,000-175,000, 95,000, and 75,000. The formation of these complexes was competitively inhibited by unlabeled TNF. Partial reversal of cross-linking in these complexes and their analysis by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) resolved 125I-TNF dimers cleaved from the 95,000 band and 125I-TNF monomer cleaved from the 75,000 band, providing evidence for a Mr approximately 60,000 subunit. In addition, the 95,000 and 75,000 bands were resolved as components of larger complexes (Mr = 150,000-175,000), which presumably contain two receptor subunits. The Mr 95,000 and 75,000 bands were also released from the Mr 310,000 complex by reduction with dithiothreitol, suggesting a role for disulfide bond stabilization. To investigate the association of the putative receptor subunits, Triton X-100 extracts from HeLa membranes were fractionated by SDS-PAGE without reduction and transferred electrophoretically to nylon membranes for TNF binding assays. Only two bands of Mr = 60,000 and 70,000 specifically bound TNF, and higher Mr binding activity was not observed. These results indicate that TNF receptors in HeLa cells are high molecular weight complexes containing Mr = 60,000 and 70,000 subunits each capable of binding TNF and that the complexes are primarily stabilized by non-covalent, hydrophobic interactions.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
5
pubmed:volume
264
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
14646-52
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1989
pubmed:articleTitle
Multimeric structure of the tumor necrosis factor receptor of HeLa cells.
pubmed:affiliation
Department of Biological Sciences, State University of New York, Albany 12222.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't