Henley J M, Nielsen M, Barnard E A
Department of Pharmacology, Medical School, University of Birmingham, England.
J Neurochem. 1992 Jun;58(6):2030-6. doi: 10.1111/j.1471-4159.1992.tb10943.x.
alpha-[3H]Amino-3-hydroxy-5-methylisoxazolepropionate ([3H]AMPA) binds to 1-day-old chick telencephalon membranes with KD and Bmax values of 138 nM and 2.56 pmol/mg of protein, respectively. High-energy radiation bombardment of intact frozen telencephalon resulted in a biphasic inactivation curve for [3H]AMPA binding. At a 5.8-Mrad radiation dose, the affinity of [3H]AMPA binding was increased (54 nM), but there was no apparent alteration in the Bmax value (2.76 pmol/mg of protein). We attribute this phenomenon to the inactivation of a high molecular weight modulatory protein that down-regulates the affinity of [3H]AMPA binding. The estimated molecular masses of the AMPA binding site and of the modulatory component were 59 and 108 kDa, respectively. Solubilisation with n-octyl-beta-glucopyranoside resulted in an increase in the Bmax (4.7 pmol/mg of protein) with no pronounced alteration in the affinity (109 nM) of [3H]AMPA binding. However, the solubilisation-induced increase in Bmax did not occur in telencephalon irradiated before solubilisation. In contrast, the increase in affinity induced by radiation treatment was still detected in solubilised extracts. These results suggest that the number and affinity of [3H]AMPA sites in chick telencephalon are closely regulated and that the modulatory systems involved are affected by both irradiation and solubilisation.
α-[³H]氨基-3-羟基-5-甲基异恶唑丙酸酯（[³H]AMPA）与1日龄雏鸡端脑细胞膜结合，其解离常数（KD）和最大结合容量（Bmax）值分别为138 nM和2.56 pmol/mg蛋白质。对完整冷冻端脑进行高能辐射轰击，导致[³H]AMPA结合出现双相失活曲线。在5.8兆拉德的辐射剂量下，[³H]AMPA结合的亲和力增加（54 nM），但Bmax值无明显变化（2.76 pmol/mg蛋白质）。我们将此现象归因于一种高分子量调节蛋白的失活，该蛋白下调[³H]AMPA结合的亲和力。AMPA结合位点和调节成分的估计分子量分别为59 kDa和108 kDa。用正辛基-β-D-吡喃葡萄糖苷增溶导致Bmax增加（4.7 pmol/mg蛋白质），而[³H]AMPA结合的亲和力无明显改变（109 nM）。然而，增溶诱导的Bmax增加在增溶前照射的端脑中未出现。相反，在增溶提取物中仍能检测到辐射处理诱导的亲和力增加。这些结果表明，雏鸡端脑中[³H]AMPA位点的数量和亲和力受到密切调节，且所涉及的调节系统受辐射和增溶的影响。
