The latency of rat liver microsomal protein disulphide-isomerase.
The latency of rat liver microsomal protein disulphide-isomerase.
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大鼠肝微粒体蛋白二硫键异构酶的潜伏期。
DOI：
10.1042/bj2280635
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发表时间：
1985
期刊：
影响因子：
--
通讯作者：
R. Freedman
中科院分区：
文献类型：
--
作者：
N. Lambert;R. Freedman
在新鲜制备的大鼠肝微粒体（微粒体部分）中未检测到蛋白二硫化物异构酶（PDI）活性，但在损伤膜完整性的处理（例如超声处理、去污剂处理或冻融）后可检测到。超声处理后可检测到最大活性。在通过凝胶过滤制备的微粒体和通过高速离心制备的微粒体中观察到相同的潜伏期。PDI活性在所有颗粒亚细胞组分中是潜伏的，但在高速上清液中不是潜伏的。当所有的馏分进行超声处理，暴露总PDI活性，PDI被发现在微粒体馏分中的最高比活性和共分布的内质网的标记酶。在各种条件下洗涤微粒体，去除外周蛋白质，在某些情况下，结合核糖体没有去除大量的PDI，也没有影响PDI活性的潜伏期。在微粒体囊泡的渗透屏障保持完整的条件下，用蛋白酶处理微粒体并没有显著抑制PDI或影响其潜伏期。PDI是非常容易从微粒体囊泡溶解的低浓度的洗涤剂，其中除去只有一小部分的总微粒体蛋白。在所有这些方面，PDI类似于核苷二磷酸酶，一种内质网腔表面的标记外周蛋白，并且不同于NADPH：细胞色素c还原酶，一种暴露在膜的细胞质表面的标记整合蛋白。这些数据是兼容的模型，其中PDI是松散的与腔面的内质网，位置与建议的生理作用的酶作为催化剂的天然二硫键的形成在新生和新合成的分泌蛋白相一致。
Protein disulphide-isomerase (PDI) activity was not detectable in freshly prepared rat liver microsomes (microsomal fraction), but became detectable after treatments that damage membrane integrity, e.g. sonication, detergent treatment or freezing and thawing. Maximum activity was detectable after sonication. Identical latency was observed in microsomes prepared by gel filtration and in those prepared by high-speed centrifugation. PDI activity was latent in all particulate subcellular fractions, but not latent in the high-speed supernatant. When all fractions were sonicated to expose total PDI activity, PDI was found at highest specific activity in the microsomal fraction and co-distributed with marker enzymes of the endoplasmic reticulum. Washing of microsomes under various conditions that removed peripheral proteins and, in some cases, bound ribosomes did not remove significant quantities of PDI, nor did it affect the latency of PDI activity. Treatment of microsomes with proteinases, under conditions where the permeability barrier of the microsomal vesicles was maintained intact, did not inactivate PDI significantly or affect its latency. PDI was very readily solubilized from microsomal vesicles by low concentrations of detergents, which removed only a fraction of the total microsomal protein. In all these respects, PDI resembled nucleoside diphosphatase, a marker peripheral protein of the luminal surface of the endoplasmic reticulum, and differed from NADPH: cytochrome c reductase, a marker integral protein exposed at the cytoplasmic surface of the membrane. The data are compatible with a model in which PDI is loosely associated with the luminal surface of the endoplasmic reticulum, a location consistent with the proposed physiological role of the enzyme as catalyst of formation of native disulphide bonds in nascent and newly synthesized secretory proteins.