Interaction of calreticulin with protein disulfide isomerase
Interaction of calreticulin with protein disulfide isomerase
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DOI：
10.1074/jbc.270.52.31338
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发表时间：
1995-12-29
影响因子：
4.8
通讯作者：
Michalak, M
中科院分区：
文献类型：
--
作者：
Baksh, S;Burns, K;Michalak, M
本文报道钙网蛋白与蛋白质二硫键异构酶（PDI）的相互作用，PDI-钙网蛋白复合物可被Zn ~（2+）-亚氨基二乙酸取代琼脂糖凝胶层析分离，表明这种相互作用可能是Zn ~（2+）依赖的。钙网蛋白和PDI之间的直接相互作用也通过钙网蛋白亲和层析记录。PDI是唯一保留在钙网蛋白亲和柱上的胰腺微粒体蛋白。钙网蛋白和PDI通过其NH 2-末端氨基酸序列分析、SDS-聚丙烯酰胺凝胶电泳迁移率、Ca-45（2+）结合以及与特异性抗体的反应性来鉴定。使用谷胱甘肽S-转移酶-钙网蛋白融合蛋白，我们表明，PDI强烈相互作用的P-结构域和N-结构域的钙网蛋白只有微弱的。在酵母双杂交系统中表达钙网蛋白结构域和PDI作为与GAL 4的融合蛋白，揭示钙网蛋白在正常细胞条件下也与PDI相互作用。与PDI的相互作用仅需要钙网蛋白的N-结构域（氨基酸残基1-83）和P-结构域（氨基酸残基150-240）的NH 2-末端区域。重要的是，钙网蛋白和PDI之间的相互作用导致其活动的调制。在存在PDI的情况下，钙网蛋白不以高亲和力结合Ca 2+。钙网蛋白或钙网蛋白的N-结构域抑制PDI重折叠乱序RNA酶A的能力。
We report here that calreticulin interacts with protein disulfide isomerase (PDI), The PDI-calreticulin complex can be dissociated by Zn2+-iminodiacetate substituted Sepharose-agarose chromatography, suggesting that these interactions may be Zn2+ dependent. Direct interaction between calreticulin and PDI is also documented by calreticulin affinity chromatography. PDI was the only pancreatic microsomal protein retained on the calreticulin affinity column. Calreticulin and PDI were identified by their NH2-terminal amino acid sequence analysis, mobilities in SDS-polyacrylamide gel electrophoresis, binding of Ca-45(2+), and their reactivity with specific antibodies. Using glutathione S-transferase-calreticulin fusion proteins, we show that PDI interacts strongly with the P-domain and only weakly with the N-domain of calreticulin. Expression of calreticulin domains and PDI as fusion proteins with GAL4 in the yeast two-hybrid system revealed that calreticulin interacted with PDI also under normal cellular conditions. Interaction with PDI required only the NH2-terminal region of the N-domain (amino acid residues 1-83) and the P-domain (amino acid residues 150-240) of calreticulin. Importantly, interaction between calreticulin and PDI led to the modulation of their activities. In the presence of PDI, calreticulin does not bind Ca2+ with high affinity. Calreticulin or the N-domain of calreticulin inhibited PDI ability to refold scrambled RNase A.