Multimeric structure of the tumor necrosis factor receptor of HeLa cells.

摘要: Abstract The tumor necrosis factor (TNF) receptor of HeLa cells was solubilized in Triton X-100 and characterized by gel filtration, affinity labeling, ligand blotting studies. Receptors with eluted filtration as a major peak Mr = 330,000 retained high binding (KD 0.25 nM). Affinity labeling soluble receptor/125I-TNF complexes using the reversible, bifunctional bis[2-(succinimidooxycarbonyl-oxy)ethyl] sulfone resulted formation cross-linked species 310,000, 150,000-175,000, 95,000, 75,000. these competitively inhibited unlabeled TNF. Partial reversal cross-linking their analysis two-dimensional sodium dodecyl sulfate-polyacrylamide electrophoresis (SDS-PAGE) resolved 125I-TNF dimers cleaved from 95,000 band monomer 75,000 band, providing evidence for approximately 60,000 subunit. In addition, bands were components larger (Mr 150,000-175,000), which presumably contain two subunits. also released 310,000 complex reduction dithiothreitol, suggesting role disulfide bond stabilization. To investigate association putative subunits, extracts membranes fractionated SDS-PAGE without transferred electrophoretically to nylon TNF assays. Only 70,000 specifically bound TNF, higher activity not observed. These results indicate that receptors are molecular weight containing subunits each capable primarily stabilized non-covalent, hydrophobic interactions.