COMMON PRECURSOR TO CORTICOTROPINS AND ENDORPHINS
COMMON PRECURSOR TO CORTICOTROPINS AND ENDORPHINS
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DOI：
10.1073/pnas.74.7.3014
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发表时间：
1977-01-01
影响因子：
11.1
通讯作者：
LING, N
中科院分区：
文献类型：
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作者：
MAINS, RE;EIPPER, BA;LING, N
使用内啡肽和促肾上腺皮质激素 (ACTH) 抗血清的双抗体免疫沉淀程序来研究这些肽在小鼠垂体肿瘤细胞系中的生物合成。将培养物与 3H 标记的氨基酸一起孵育，并对等份培养基进行免疫沉淀。使用内啡肽抗血清制备的[3H]苯丙氨酸标记的免疫沉淀物进行十二烷基硫酸钠/聚丙烯酰胺凝胶电泳，分离出 3 种形式的内啡肽，表观分子量分别为 31,000、11,700 和 3500；用 ACTH 抗血清制备的免疫沉淀物含有 4 种形式的 ACTH，表观分子量分别为 31,000、23,000、13,000 和 < 4500。用 ACTH 抗血清然后用内啡肽抗血清（或相反的顺序）对培养基进行连续免疫沉淀表明两种抗血清沉淀了相同的 31,000 道尔顿分子。通过免疫沉淀和凝胶过滤制备不同形式的 ACTH 和内啡肽的纯化池。 [3H]苯丙氨酸或[3H]色氨酸标记的 31,000 道尔顿 ACTH 中发现的胰蛋白酶肽与用相同氨基酸标记的 31,000 道尔顿内啡肽消化物中发现的胰蛋白酶肽相同。可以在 31,000 道尔顿的 ACTH 和所有不同形式的内啡肽中鉴定出与促脂蛋白胰蛋白酶肽 [βLPH(61-69)] 相似的胰蛋白酶肽，其含有阿片活性甲硫氨酸-脑啡肽序列。当 31,000 道尔顿 ACTH 转化为 23,000 道尔顿 ACTH 时，大部分肽被裂解，可被内啡肽抗血清沉淀；这种 11,700 道尔顿的内啡肽分子在大小和结构上与垂体激素 .beta.LPH 相似。 3500道尔顿的内啡肽在大小和结构上与β-内啡肽相似。 AtT-20小鼠垂体肿瘤细胞的培养基含有大约等摩尔量的ACTH相关肽和内啡肽相关肽。
Double-antibody immunoprecipitation procedures with antisera to endorphins and to corticotropin (ACTH) were used to study the biosynthesis of these peptides in a mouse pituitary tumor cell line. Cultures were incubated with a 3H-labeled amino acid, and aliquots of culture medium were immunoprecipitated. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis of [3H]phenylalanine-labeled immunoprecipitates prepared with endorphin antisera resolved 3 forms of endorphin with apparent MW of 31,000, 11,700, and 3500; immunoprecipitates prepared with the ACTH antiserum contained 4 forms of ACTH with apparent MW of 31,000, 23,000, 13,000, and < 4500. Sequential immunoprecipitation of culture medium with the ACTH antiserum and then with the endorphin antiserum (or the reverse order) indicated that both antisera precipitated the same 31,000 dalton molecule. Purified pools of the different forms of ACTH and endorphin were prepared by immunoprecipitation and gel filtration. The tryptic peptides found in [3H]phenylalanine- or [3H]tryptophan-labeled 31,000 dalton ACTH were identical to the tryptic peptides found in digests of 31,000 dalton endorphin labeled with the same amino acid. A tryptic peptide similar to the lipotropin tryptic peptide [.beta.LPH(61-69)] that contains the opiate-active methionine-enkephalin sequence could be identified in 31,000 dalton ACTH and in all the different forms of endorphin. Most of the peptide cleaved from 31,000 dalton ACTH when it is converted to 23,000 dalton ACTH could be precipitated by endorphin antisera; this 11,700 dalton endorphin molecule is similar to the pituitary hormone .beta.LPH in size and structure. The 3500 dalton endorphin is similar to .beta.-endorphin in size and structure. The culture medium from the AtT-20 mouse pituitary tumor cells contained approximately equimolar amounts of ACTH-related peptides and endorphin-related peptides.