Lee T H, Lee M S
Biochemistry. 1977 Jun 28;16(13):2824-9. doi: 10.1021/bi00632a003.
A highly purified preparation of high-molecular-weight adrenocorticotropic hormone (ACTH) was prepared from ovine pituitary glands by dilute acetic acid extraction, oxycellulose fractionation. Sephadex gel filtration, and affinity chromatography on immobilized alphap(1-39)ACTH antibodies. Two ACTH peptides of molecular weights of 24 000 and 34 000 were detected by sodium dodecyl sulfate-acrylamide gel electrophoresis in this preparation. It appeared that the immobilized antibodies adsorbed two forms equally well and could not distinguish between them under the conditions used. These two ACTH peptides were found to be present in crude extracts of ovine pituitary glands, indicating that they were not artifacts produced by the purification procedure. The high-molecular-weight forms of ACTH were found to be susceptible to degradation by tissue enzymes. They could be easily destroyed during the extraction, if precautions were not taken. Moreover, they were poorly adsorbed by oxycellulose which had been used for the adsorption of ACTH activity from crude preparations by most investigators. These properties probably accounted for the fact that high-molecular-weight forms of ACTH remained undetected until very recently.
通过稀醋酸萃取、羟甲基纤维素分级分离、葡聚糖凝胶过滤以及在固定化α(1-39)促肾上腺皮质激素抗体上进行亲和层析，从绵羊脑垂体中制备了一种高度纯化的高分子量促肾上腺皮质激素（ACTH）制剂。在该制剂中，通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳检测到分子量分别为24000和34000的两种促肾上腺皮质激素肽。看来固定化抗体对这两种形式的吸附效果相同，在所使用的条件下无法区分它们。发现这两种促肾上腺皮质激素肽存在于绵羊脑垂体的粗提物中，这表明它们不是纯化过程产生的假象。已发现高分子量形式的促肾上腺皮质激素易被组织酶降解。如果不采取预防措施，它们在提取过程中很容易被破坏。此外，它们被大多数研究人员用于从粗制品中吸附促肾上腺皮质激素活性的羟甲基纤维素吸附得很差。这些特性可能解释了直到最近才检测到高分子量形式的促肾上腺皮质激素这一事实。
