Duan D S, Pazin M J, Fretto L J, Williams L T
Cardiovascular Research Institute, Howard Hughes Medical Institute, University of California, San Francisco 94143.
J Biol Chem. 1991 Jan 5;266(1):413-8.
The platelet-derived growth factor beta-receptor (PDGFr) is a 180-kDa transmembrane glycoprotein which binds BB-PDGF with high affinity. We have expressed the extracellular region of the receptor in Chinese hamster ovary cells using an expression vector that carries a dihydrofolate reductase gene as an amplifiable marker. Upon amplification of the receptor cDNA sequences by methotrexate a 110-kDa soluble form of the receptor extracellular region (XR) was secreted at 12 mg/liter. The soluble XR protein fully retained the high affinity specific binding of the intact PDGFr for BB-PDGF (apparent dissociation constant, 0.4 nM). In the presence of ligand the soluble XR protein formed complexes that migrated on sodium dodecyl sulfate gels at the size expected for dimers of the protein. When added to fibroblast cultures the soluble XR protein blocked the ability of BB-PDGF to stimulate DNA synthesis but did not alter the mitogenic effect of AA-PDGF. The XR fragment also inhibited the binding of BB-PDGF to PDGFr and the activation of PDGFr tyrosine kinase by BB-PDGF. Thus, the soluble extracellular region protein of the PDGFr binds BB-PDGF with high affinity and functions as a specific antagonist of BB-PDGF actions.
血小板衍生生长因子β受体（PDGFr）是一种180 kDa的跨膜糖蛋白，它能与BB-PDGF高亲和力结合。我们利用携带二氢叶酸还原酶基因作为可扩增标记的表达载体，在中国仓鼠卵巢细胞中表达了该受体的细胞外区域。通过甲氨蝶呤扩增受体cDNA序列后，一种110 kDa的受体细胞外区域可溶性形式（XR）以12 mg/升的浓度分泌出来。可溶性XR蛋白完全保留了完整PDGFr对BB-PDGF的高亲和力特异性结合（表观解离常数为0.4 nM）。在有配体存在的情况下，可溶性XR蛋白形成的复合物在十二烷基硫酸钠凝胶上的迁移大小符合该蛋白二聚体的预期大小。当添加到成纤维细胞培养物中时，可溶性XR蛋白阻断了BB-PDGF刺激DNA合成的能力，但不改变AA-PDGF的促有丝分裂作用。XR片段还抑制了BB-PDGF与PDGFr的结合以及BB-PDGF对PDGFr酪氨酸激酶的激活。因此，PDGFr的可溶性细胞外区域蛋白能与BB-PDGF高亲和力结合，并作为BB-PDGF作用的特异性拮抗剂发挥作用。
