Pomés A, Pong S S, Schaeffer J M
Merck Research Laboratories, Rahway, New Jersey 07065, USA.
Biochem Biophys Res Commun. 1996 Aug 23;225(3):939-45. doi: 10.1006/bbrc.1996.1275.
The discovery of a potential new GH therapy by small molecules that induce GH secretion (GHRP-6, L-692,429, MK-0677), has increased the interest in these GH secretagogues and their receptor and mechanism of action, which is different from the one of GHRH. We report the solubilization of the GH-secretagogue-receptor-ligand-G-protein complex (apparent molecular mass of approximately 255 kDa) from porcine anterior pituitary membranes using digitonin, after labelling the receptor with [35S]MK-0677. The solubilized receptor showed high affinity (KD = 122.2 +/- 14.4 pM) and low capacity (Bmax = 3.8 +/- 0.9 fmol/mg protein). These values and the inhibition constants (Ki) for a series of GH secretagogues were similar to the values determined in membranes isolated from porcine anterior pituitary gland. The solubilization of the GH secretagogue receptor opens up the possibility for further molecular characterization and sequencing of the receptor protein, necessary step prior to the identification of the natural ligand that would act as a GHRH amplifying hormone, and that the GH secretagogues would mimic.
通过诱导生长激素（GH）分泌的小分子（生长激素释放肽-6、L-692,429、MK-0677）发现了一种潜在的新型GH疗法，这增加了人们对这些GH促分泌素及其受体和作用机制的兴趣，其作用机制与生长激素释放激素（GHRH）不同。我们报告了在用[35S]MK-0677标记受体后，使用洋地黄皂苷从猪垂体前叶膜中溶解GH促分泌素受体-配体-G蛋白复合物（表观分子量约为255 kDa）。溶解的受体显示出高亲和力（KD = 122.2 +/- 14.4 pM）和低容量（Bmax = 3.8 +/- 0.9 fmol/mg蛋白）。这些值以及一系列GH促分泌素的抑制常数（Ki）与从猪垂体前叶分离的膜中测定的值相似。GH促分泌素受体的溶解为受体蛋白的进一步分子表征和测序开辟了可能性，这是鉴定作为GHRH放大激素的天然配体以及GH促分泌素模拟物的必要步骤。
