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首页> 外文期刊>Biochemistry >Transfected cells express mostly the intracellular precursor of the lutropin/choriogonadotropin receptor but this precursor binds choriogonadotropin with high affinity
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Transfected cells express mostly the intracellular precursor of the lutropin/choriogonadotropin receptor but this precursor binds choriogonadotropin with high affinity

机译：转染的细胞主要表达促性腺激素/绒毛膜促性腺激素受体的细胞内前体，但该前体以高亲和力结合绒毛膜促性腺激素

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Previous studies from several laboratories have shown that the cell surface rLHR is a 85-92 kDa protein synthesized from a 68-73 kDa intracellular precursor. While all investigators agree that the cell surface rLHR binds hCG with high affinity, it is not clear if the intracellular precursor can also bind hCG. In order to directly determine if the intracellular rLHR present in cells transfected with the wild-type rLHR binds hCG with high affinity, we devised a method that selectively degrades the cell surface rLHR while preserving the intracellular rLHR. The binding of hCG to intact cells was completely lost following mild proteolysis of The cells, but binding to detergent extracts prepared from proteolyzed cells was largely preserved. Measurements of the hCG binding affinity to intact cells or to detergent extracts prepared before and after proteolysis display very similar or identical binding affinities. Since binding to nonproteolyzed intact cells, detergent extracts prepared from nonproteolyzed cells, or detergent extracts prepared from proteolyzed cells occurs only to the 85-92 kDa rLHR, the 85-92 and 68-73 MDa rLHR, and the 68-73 kDa rLHR, respectively, we conclude that the cell surface rLHR and the intracellular rLHR bind hCG with the same affinity. Quantitation of the relative abundance: of the cell surface and intracellular rLHR by immunological methods indicates that transfected cells express mostly the intracellular precursor. A comparison of the binding capacity of control and proteolyzed cells with that of their detergent extracts indicates that hCG binding assays greatly underestimate the relative abundance of the intracellular rLHR. [References: 33]
机译：来自多个实验室的先前研究表明，细胞表面rLHR是由68-73 kDa细胞内前体合成的85-92 k​​Da蛋白。尽管所有研究人员都认为细胞表面rLHR以高亲和力结合hCG，但尚不清楚细胞内前体是否也可以结合hCG。为了直接确定用野生型rLHR转染的细胞中存在的细胞内rLHR是否以高亲和力结合hCG，我们设计了一种在保留细胞内rLHR的同时选择性降解细胞表面rLHR的方法。在温和地对细胞进行蛋白水解后，hCG与完整细胞的结合完全消失，但与蛋白水解细胞制备的去污剂提取物的结合得以保留。对蛋白水解前后制备的完整细胞或去污剂提取物的hCG结合亲和力的测量显示出非常相似或相同的结合亲和力。由于与非蛋白水解完整细胞结合，因此从非蛋白水解细胞制备的去污剂提取物或从蛋白水解细胞制备的去污剂提取物仅在85-92 k​​Da rLHR，85-92和68-73 MDa rLHR以及68-73 kDa rLHR处发生，我们分别得出结论，细胞表面rLHR和细胞内rLHR以相同的亲和力结合hCG。通过免疫学方法对细胞表面和细胞内rLHR的相对丰度进行定量分析，表明转染的细胞主要表达细胞内前体。对照细胞和蛋白水解细胞与其去污剂提取物的结合能力的比较表明，hCG结合测定法大大低估了细胞内rLHR的相对丰度。 [参考：33]

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