Coffer A, Fabregat I, Sinnett-Smith J, Rozengurt E
Imperial Cancer Research Fund, Lincoln's Inn Fields, London, UK.
FEBS Lett. 1990 Apr 9;263(1):80-4. doi: 10.1016/0014-5793(90)80710-z.
Bombesin and structurally related peptides including gastrin releasing peptide (GRP) are potent mitogens for Swiss 3T3 cells. Here we attempted to solubilize bombesin receptors under conditions in which the ligand (125I-labelled GRP) was prebound to the receptor prior to detergent extraction. We found that 125I-GRP-receptor complexes were solubilized from Swiss 3T3 cell membranes by using the detergents taurodeoxycholate or deoxycholate. These detergents promoted ligand-receptor solubilization in a dose-dependent manner. In contrast, a variety of other detergents including Triton X-100, octylglycoside, CHAPS, digitonin, cholic acid and n-dodecyl-beta-D-maltoside, were much less effective. Addition of guanosine 5'-[gamma-thio]triphosphate (GTP gamma S) to ligand-receptor complexes isolated by gel filtration enhanced the rate of ligand dissociation in a concentration-dependent and nucleotide-specific manner. Our results demonstrate for the first time the successful solubilization of 125I-GRP-receptor complexes from Swiss 3T3 cell membranes and provide evidence for the physical association between the ligand-receptor complex and a guanine nucleotide binding protein(s).
蛙皮素及包括胃泌素释放肽（GRP）在内的结构相关肽是瑞士3T3细胞的强效促细胞分裂剂。在此，我们尝试在去污剂提取之前使配体（125I标记的GRP）预先结合到受体的条件下溶解蛙皮素受体。我们发现，使用牛磺去氧胆酸盐或去氧胆酸盐去污剂可从瑞士3T3细胞膜中溶解125I-GRP-受体复合物。这些去污剂以剂量依赖的方式促进配体-受体的溶解。相比之下，包括Triton X-100、辛基糖苷、CHAPS、洋地黄皂苷、胆酸和正十二烷基-β-D-麦芽糖苷在内的多种其他去污剂效果要差得多。向通过凝胶过滤分离的配体-受体复合物中添加鸟苷5'-[γ-硫代]三磷酸（GTPγS），以浓度依赖和核苷酸特异性的方式提高了配体解离的速率。我们的结果首次证明了从瑞士3T3细胞膜中成功溶解125I-GRP-受体复合物，并为配体-受体复合物与鸟嘌呤核苷酸结合蛋白之间的物理关联提供了证据。
