Purification of a 130-kDa T cell glycoprotein that binds human interleukin 4 with high affinity.

作者：Jean‐Pierre Galizzi, Brian E. Castle, Odile Djossou, Nobuyuki Harada, H Cabrillat, Saliha Ait Yahia, Rachel L. C. Barrett, M Howard, Jacques Banchereau · 发表于：Journal of Biological Chemistry · 年份：1990 · DOI：10.1016/s0021-9258(19)40249-4 · 被引用次数：42 · 研究领域：Monoclonal and Polyclonal Antibodies Research、Toxin Mechanisms and Immunotoxins、T-cell and B-cell Immunology

The interleukin 4 (IL-4) receptor was purified from the gibbon T cell line MLA 144. These cells were found to express high numbers of human IL-4-binding proteins (5000-6000 sites/cell) with an affinity constant (Kd) similar to that measured in human cell lines (Kd = 40-70 pM). Affinity cross-linking of 125I-IL-4 to human cell lines and MLA 144 cells demonstrated the labeling of three proteins of approximately 130, 75, and 65 kDa. Human IL-4-binding sites were solubilized from MLA 144 cells using Triton X-100 and then purified by carboxymethyl chromatography, which removed 50% of the protein without loss of IL-4-binding activity. Then sequential affinity purification over wheat germ agglutinin and a single IL-4 Affi-Gel 10 column resulted in a final 8000-fold purification of the IL-4 receptor. When analyzed on a silver-stained sodium dodecyl sulfate-polyacrylamide gel, the purified receptor migrated as a single molecular species of 130 +/- 5 kDa. Identification of the 130-kDa protein as the IL-4 receptor was demonstrated by cross-linking experiments and specific binding of 125I-IL-4 to nitrocellulose membranes after electrophoretic transfer of the purified receptor on sodium dodecyl sulfate-polyacrylamide gel.