Identification of cysteine-rich domains of the type 1 tumor necrosis factor receptor involved in ligand binding.

摘要: The extracellular portion of the type 1 (p55) and 2 (p75) tumor necrosis factor (TNF) receptors contains a repetitive amino acid sequence pattern four cysteine-rich domains (CRDs). This is found also in several other cell surface proteins, including p75 nerve growth receptor CD40, 4-1BB, OX40, Fas, CD27 antigens. To investigate whether CRDs play role TNF binding, we have constructed soluble variants human (sTNFR1), which CRD1 (N-terminal) or CRD4 (C-terminal) was deleted by mutagenesis. These wild sTNFR1 were linked their C terminus to hinge Fc IgG1 heavy chain create sTNFR1-IgG chimeras (immunoadhesins). Deletion either -4 did not cause any major perturbations structure variants, as evidenced efficient expression secretion from transfected cells, binding conformation-dependent monoclonal antibodies that recognize diverse epitopes on sTNFR1. immunoadhesin exhibited high affinity alpha (Kd = 65 pM) beta 640 pM). resulted about 10-fold reduction for 660 5.7 nM). In contrast, deletion complete loss beta. results indicate important but necessary while required. addition, suggest some overlap between TNFR1 sites beta, despite low homology these cytokines.