Biochemical investigation of the subcellular localization of the estrogen-induced pro-glandular kallikrein in the rat anterior pituitary.
Biochemical investigation of the subcellular localization of the estrogen-induced pro-glandular kallikrein in the rat anterior pituitary.
复制标题
大鼠垂体前叶雌激素诱导的前腺激肽释放酶亚细胞定位的生化研究。
DOI：
10.1159/000125164
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发表时间：
1989
影响因子：
4.1
通讯作者：
Powers,CA
中科院分区：
文献类型：
--
作者：
Hatala,MA;Powers,CA
腺激肽释放酶（胰蛋白酶样丝氨酸蛋白酶）是大鼠垂体前叶中一种主要的雌激素诱导和多巴胺抑制蛋白，似乎与催乳素相关。在脑垂体中，这种酶主要以潜在酶原（前腺激肽释放酶）的形式存在，可被胰蛋白酶激活。本研究报告实验采用生化技术，调查腺激肽释放酶的亚细胞定位。从雌激素处理的大鼠垂体前叶分馏上的不连续蔗糖密度梯度和梯度中的各种细胞器的分布，通过常规的酶或蛋白质标记物测定。8个细胞器标记物中的每一个在梯度内表现出独特的分布曲线。腺激肽释放酶的分布密切相关（r = 0.91）与核苷二磷酸酶（高尔基体跨池的标志物）。对于腺激肽释放酶和核苷二磷酸酶，35-45%的总活性被发现在高尔基带的梯度，和18-22%是在分泌囊泡部分。在所有的亚细胞级分中，91-97%的腺激肽释放酶以酶原形式（前腺激肽释放酶）存在。在高尔基体组分中，38%的腺激肽释放酶在冻融和低渗介质中两次洗涤后保持膜结合; 94%的核苷二磷酸酶在这样的处理后保持膜结合。结果表明，腺激肽释放酶是最高度集中在高尔基体的transcisterebrae与大量的活动也存在于分泌囊泡。这种定位与腺激肽释放酶在催乳素细胞中作为激素原加工酶的作用一致。
Glandular kallikrein (a trypsin-like serine protease) is a major estrogen-induced and dopamine-repressed protein in the rat anterior pituitary which appears to be associated with lactotrophs. In the pituitary the enzyme predominantly exists as a latent zymogen (pro-glandular kallikrein) which can be activated by trypsin. This study reports experiments employing biochemical techniques to investigate the subcellular localization of glandular kallikrein. Anterior pituitaries from estrogen-treated rats were fractionated on a discontinuous sucrose density gradient and the distribution of various organelles in the gradient was determined by conventional enzyme or protein marker assays. Each of the 8 organelle markers exhibited a unique distribution profile within the gradient. The distribution of glandular kallikrein was closely correlated (r = 0.91) with that of nucleoside diphosphatase (a marker for trans cisterna of the Golgi apparatus). For both glandular kallikrein and nucleoside diphosphatase, 35–45% of the total activity was found in Golgi zones of the gradient, and 18–22% was in the secretory vesicle fraction. In all of the subcellular fractions, 91–97% of the glandular kallikren existed in the zymogen form (pro-glandular kallikrein). In Golgi fractions, 38% of the glandular kallikrein remained membrane-bound following freeze-thawing and two washes in hypotonic media; 94% of the nucleoside disphosphatase remained membrane-bound following such treatment. The results indicate that glandular kallikrein is most highly concentrated in trans cisternae of the Golgi apparatus with substantial activity also present in secretory vesicles. This localization is consistent with a role for glandular kallikrein as a prohormone processing enzyme in lactotrophs.