Demoliou-Mason C, Epand R M
Biochemistry. 1982 Apr 27;21(9):1989-96. doi: 10.1021/bi00538a003.
The concentration-dependent stimulation of adenylate cyclase by the photoaffinity reagent 2-[(2-nitro-4-azidophenyl)sulfenyl]-Trp25-glucagon (glucagon-NAPS) and also its binding characteristics were compared with those of the native hormone. The derivative was found to be slightly more potent in stimulating adenylate cyclase than glucagon, in the presence of guanosine 5'-triphosphate (GTP). 125I-Labeled glucagon-NAPS or 125I-labeled glucagon bound rapidly to receptors and was competitively displaced by unlabeled glucagon or glucagon-NAPS. Glucagon-NAPS displaced bound radiolabeled hormone at a lower concentration than did glucagon in the absence of GTP. Scatchard analysis of the binding data obtained from displacement of bound radiolabeled ligand with unlabeled peptide demonstrated a heterogeneous population of saturable glucagon binding sites. Glucagon-NAPS displayed a higher affinity (0.7 nM) for the high-capacity sites (80-90% of total binding sites) than glucagon (4.0 nM) in the absence of GTP. In the presence of the nucleotide, both ligands had approximately the same affinity (0.5-0.6 nM). Hill plot analysis of the binding data suggested noncooperative interactions. Photoaffinity labeling of plasma membranes with glucagon-NAPS resulted in an irreversible activation of adenylate cyclase with a reduced response to further stimulation by glucagon, glucagon-NAPS, and NaF.
将光亲和试剂2-[(2-硝基-4-叠氮苯基)亚磺酰基]-色氨酸25-胰高血糖素（胰高血糖素-NAPS）对腺苷酸环化酶的浓度依赖性刺激及其结合特性与天然激素进行了比较。发现在存在鸟苷5'-三磷酸（GTP）的情况下，该衍生物刺激腺苷酸环化酶的效力比胰高血糖素略强。125I标记的胰高血糖素-NAPS或125I标记的胰高血糖素能迅速与受体结合，并被未标记的胰高血糖素或胰高血糖素-NAPS竞争性取代。在不存在GTP的情况下，胰高血糖素-NAPS以比胰高血糖素更低的浓度取代结合的放射性标记激素。用未标记的肽取代结合的放射性标记配体所获得的结合数据的Scatchard分析表明，存在可饱和的胰高血糖素结合位点的异质群体。在不存在GTP的情况下，胰高血糖素-NAPS对高容量位点（占总结合位点的80-90%）的亲和力（0.7 nM）高于胰高血糖素（4.0 nM）。在存在核苷酸的情况下，两种配体具有大致相同的亲和力（0.5-0.6 nM）。结合数据的Hill图分析表明存在非协同相互作用。用胰高血糖素-NAPS对质膜进行光亲和标记导致腺苷酸环化酶的不可逆激活，对胰高血糖素、胰高血糖素-NAPS和NaF的进一步刺激反应降低。
