TI  - Biochemical properties of the 75-kDa tumor necrosis factor receptor.
      Characterization of ligand binding, internalization, and receptor
      phosphorylation.
PG  - 21172-8
AB  - An expression plasmid encoding the human 75-kDa <prot>tumor necrosis factor
      (TNF) type 2 receptor</prot> (<prot>TNF-R2</prot>) was constructed and used to generate a
      stable human cell line (293/<prot>TNF-R2</prot>) overexpressing   <prot>TNF-R2</prot>  . Ligand binding
      analysis revealed high affinity binding (Kd = 0.2 nM) with approximately
      94,000 +/- 7,500 sites/cell for 125I- <prot>TNF-alpha</prot>  and approximately 5-fold
      lower affinity for  <prot>TNF-beta</prot>  (Kd = 1.1 nM) with 264,000 +/- 2,000
      sites/cell. Cross-linking of 125I-<prot>TNF-alpha</prot> and 125I-<prot>TNF-beta</prot> to
      293/<prot>TNF-R2</prot> cells yielded predominant complexes with apparent molecular
      weights of 211,000 for <prot>TNF-alpha</prot> and 205,000 and 244,000 for <prot>TNF-beta</prot>,
      suggesting these complexes contain two or three <prot>TNF-R2</prot> molecules.
      Immunoprecipitation of <prot>TNF-R2</prot> from 32P-labeled 293/<prot>TNF-R2</prot> cells
      demonstrated that the receptor is phosphorylated. The majority (97%) of
      32Pi incorporation was found in serine residues with a very low level of
      incorporation (3%) in threonine residues. <prot>TNF-alpha</prot> treatment of
      293/<prot>TNF-R2</prot> cells did not significantly affect the degree or pattern of
      phosphorylation. Cell surface-bound 125I-TNF-alpha was slowly internalized
      by the 293/<prot>TNF-R2</prot> cell line with a t1/2 = 25 min. Shedding of the
      extracellular domain of <prot>TNF-R2</prot> was induced by 4 beta-phorbol 12-myristate
      13-acetate but not by <prot>TNF-alpha</prot> or <prot>TNF-beta</prot>.
AD  - Department of Molecular Biology, Genentech, Incorporated, South San
