Tan J C, Braun S, Rong H, DiGiacomo R, Dolphin E, Baldwin S, Narula S K, Zavodny P J, Chou C C
Schering-Plough Research Institute, Kenilworth, New Jersey 07033-0530, USA.
J Biol Chem. 1995 May 26;270(21):12906-11. doi: 10.1074/jbc.270.21.12906.
The extracellular region of the human interleukin-10 (hIL-10) receptor was expressed using a myeloma cell line and was purified to homogeneity by ligand-affinity chromatography. SDS-polyacrylamide gel electrophoresis analysis indicated that the soluble receptor is glycosylated and has an apparent molecular mass of 35,000-45,000. Under native conditions, soluble hIL-10 receptor was determined by gel filtration to be a monomeric protein. Soluble hIL-10 receptor was able to inhibit the binding of 125I-hIL-10 to the full-length receptor and was able to antagonize the effect of human IL-10 in cell proliferation and cytokine synthesis inhibition. The apparent dissociation constant (Kd) of soluble hIL-10 receptor was determined to be 563 +/- 59 pM, approximately 2- to 10-fold higher than that found on intact cells (Tan, J. C., Indelicato, S. R., Narula, S. K., Zavodny, P. J., and Chou, C.-C. (1993) J. Biol. Chem. 268, 21053-21059; Liu, Y., Wei, S. H.-Y., Ho, A. S.-Y., de Waal Malefyt, R., and Moore, K. W. (1994) J. Immunol. 152, 1821-1829). When hIL-10 binds soluble hIL-10 receptor in solution, a single complex was detected by gel filtration, and the complex was found to consist of two hIL-10 dimers and four soluble receptor monomers, suggesting that hIL-10 may induce a novel mode of oligomerization of the receptor upon binding.
人白细胞介素10（hIL-10）受体的胞外区通过骨髓瘤细胞系表达，并通过配体亲和层析纯化至同质。SDS-聚丙烯酰胺凝胶电泳分析表明，可溶性受体是糖基化的，表观分子量为35,000 - 45,000。在天然条件下，通过凝胶过滤确定可溶性hIL-10受体为单体蛋白。可溶性hIL-10受体能够抑制125I-hIL-10与全长受体的结合，并能够拮抗人IL-10在细胞增殖和细胞因子合成抑制方面的作用。可溶性hIL-10受体的表观解离常数（Kd）确定为563±59 pM，比完整细胞上的解离常数高约2至10倍（Tan，J.C.，Indelicato，S.R.，Narula，S.K.，Zavodny，P.J.，和Chou，C.-C.（1993）J.Biol.Chem.268，21053 - 21059；Liu，Y.，Wei，S.H.-Y.，Ho，A.S.-Y.，de Waal Malefyt，R.，和Moore，K.W.（1994）J.Immunol.152，1821 - 1829）。当hIL-10在溶液中与可溶性hIL-10受体结合时，通过凝胶过滤检测到单一复合物，并且发现该复合物由两个hIL-10二聚体和四个可溶性受体单体组成，这表明hIL-10在结合时可能诱导受体形成一种新的寡聚化模式。
