Isolation and characterization of immunoreactive somatostatin from fish pancreatic islets.
Isolation and characterization of immunoreactive somatostatin from fish pancreatic islets.
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从鱼胰岛中分离和表征免疫反应性生长抑素。
DOI：
10.1172/jci109786
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发表时间：
1980
期刊：
影响因子：
--
通讯作者：
A. Permutt
中科院分区：
文献类型：
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作者：
H. Oyama;H. Hirsch;K. Gabbay;A. Permutt
用放射免疫法标记合成的十四肽生长抑素，在斑点叉尾（Ictaluruspunctata）的主胰岛中发现了大量的免疫反应性生长抑素。这些实验的目的是分离和表征生长抑素样物质。胰岛提取物在Bio-Gel P-30柱上层析，超过90%的免疫反应性生长抑素与至少两倍于合成十四肽生长抑素的蛋白质一起迁移。该馏分通过离子交换色谱法在羧甲基纤维素和DEAE-纤维素柱上进一步纯化。两个肽获得相同的免疫反应性，这是约25%的合成生长抑素。通过薄层电泳、pH 8.9下的聚丙烯酰胺凝胶电泳和高压液相色谱法判断每种肽的纯度> 95%。纯度的进一步标准包括仅产生天冬氨酸的级分IV的氨基末端分析。从10 g新鲜冷冻胰岛中获得总共1.3 mg级分II和3.8 mg级分IV生长抑素样肽。两种肽的表征显示两种肽比合成的十四肽生长抑素略酸性。级分II的等电点为8.0 - 8.3，级分IV的等电点为8.3 - 9.0。通过十二烷基硫酸钠-尿素聚丙烯酰胺凝胶电泳进行的分子量估计显示两种肽在胰多肽（mol wt 4，500）和胰高血糖素（mol wt 3，500）之间具有相似的迁移率。迁移率不改变还原，约为合成十四肽生长抑素（摩尔重量1，800）的两倍。这证实了肽是单一多肽链，而不是聚集体，或生长抑素结合到更大的蛋白质。通过凝胶过滤色谱法在Bio-Gel P-6上在8 M尿素中测定分子量，得到估计的摩尔重量为3，700。两种免疫反应性生长抑素的氨基酸分析表明，它们在组成上非常相似。胰腺生长抑素（1 μ M）有充分的生物活性，相对于合成生长抑素测量生长激素从大鼠垂体前叶细胞释放的抑制。
Using a radioimmunoassay with labeled synthetic tetradecapeptide somatostatin, a large amount of immunoreactive somatostatin was found in the principal pancreatic islet of the channel catfish (Ictalurus punctata). The purpose of these experiments was to isolate and characterize the somatostatin-like material. Extracts of islets were chromatographed on a Bio-Gel P-30 column, and over 90% of the immunoreactive somatostatin migrated with proteins at least twice the size of synthetic tetradecapeptide somatostatin. This fraction was further purified by ion-exchange chromatography on carboxymethyl-cellulose and DEAE-cellulose columns. Two peptides were obtained with identical immunoreactivity, which was approximately 25% that of the synthetic somatostatin. Each peptide was judged to be >95% pure by thin-layer electrophoresis, polyacrylamide gel electrophoresis at pH 8.9, and highpressure liquid chromatography. Further criteria of purity included amino-terminal analysis of fraction IV yielding only aspartic acid. A total of 1.3 mg of fraction II, and 3.8 mg of fraction IV somatostatin-like peptides were obtained from 10 g of fresh frozen islets. Characterization of the two peptides revealed both peptides slightly more acidic than synthetic tetradecapeptide somatostatin. Fraction II had an isoelectric point of 8.0-8.3, and fraction IV 8.3-9.0. Molecular weight estimation by sodium dodecyl sulfate-urea polyacrylamide gel electrophoresis revealed similar mobility of both peptides, between pancreatic polypeptide (mol wt 4,500) and glucagon (mol wt 3,500). The mobility was not altered by reduction, and was approximately twice the size of synthetic tetradecapeptide somatostatin (mol wt 1,800). This confirmed that the peptides were single polypeptide chains and not aggregates, or somatostatin bound to larger proteins. Molecular weight determination by gel filtration chromatography on Bio-Gel P-6 in 8 M urea gave an estimated mol wt of 3,700. Amino acid analysis of the two immunoreactive somatostatins indicated that they were very similar in composition. Both pancreatic somatostatins (1 muM) had full biological activity relative to synthetic somatostatin measured as inhibition of growth hormone release from rat anterior pituitary cells.