Elton T S, Oparil S, Blalock J E
Department of Medicine, University of Alabama, Birmingham 35294.
J Hypertens Suppl. 1988 Dec;6(4):S404-7. doi: 10.1097/00004872-198812040-00127.
We used the molecular recognition hypothesis, that peptide ligands and their receptor binding sites are encoded by complementary nucleotide sequences, to purify an angiotensin II (Ang II) binding protein. The complementary peptide IIA (Lys-Gly-Val-Asp-Val-Tyr-Ala-Val) specified by the RNA sequence complementary to the messenger (m)RNA sequence for rat Ang II was synthesized, purified and used to raise polyclonal antibodies. Complementary peptide IIA specifically inhibited the binding of 125I-Ang II to receptors on rat adrenal membranes, and anti-IIA immunoglobulin G (IgG) specifically inhibited the binding of 125I-Ang II to rat adrenal Ang II receptors and Ang II-dependent aldosterone secretion by cultured rat adrenal cells, suggesting that the antibody recognizes the Ang II receptor. Anti-IIA IgG was used for immuno-affinity purification, from a rat adrenal membrane preparation of an Ang II binding protein with a molecular weight of 66,000 +/- 2000 that bound 125I-Ang II specifically. This is the first report of purification of an Ang II receptor binding protein which retains its capacity to specifically bind 125I-Ang II.
我们运用分子识别假说，即肽配体及其受体结合位点由互补核苷酸序列编码，来纯化一种血管紧张素II（Ang II）结合蛋白。由与大鼠Ang II信使（m）RNA序列互补的RNA序列所指定的互补肽IIA（赖氨酸-甘氨酸-缬氨酸-天冬氨酸-缬氨酸-酪氨酸-丙氨酸-缬氨酸）被合成、纯化并用于制备多克隆抗体。互补肽IIA特异性抑制125I-Ang II与大鼠肾上腺膜上受体的结合，抗IIA免疫球蛋白G（IgG）特异性抑制125I-Ang II与大鼠肾上腺Ang II受体的结合以及培养的大鼠肾上腺细胞中Ang II依赖性醛固酮分泌，这表明该抗体识别Ang II受体。抗IIA IgG用于免疫亲和纯化，从大鼠肾上腺膜制剂中纯化出一种分子量为66,000±2000的Ang II结合蛋白，该蛋白能特异性结合125I-Ang II。这是关于纯化保留特异性结合125I-Ang II能力的Ang II受体结合蛋白的首次报道。
