Agarwal R P, McPherson R A, Threatte G A
Transplantation. 1986 Dec;42(6):627-32. doi: 10.1097/00007890-198612000-00010.
To investigate the phenomenon of different erythrocyte saturation capacities for cyclosporine (CsA) in the blood of different individuals, hemolysates of washed red cells were examined for the presence of a CsA-binding protein. Using gel filtration column chromatography of hemolysates from patients receiving CsA orally, the majority of erythrocyte-associated CsA eluted as a single peak with Mr 15,000-17,000, distinct from hemoglobin and carbonic anhydrase. [3H]CsA added to a hemolysate in vitro eluted similarly. [125I]CsA added to a hemolysate eluted much later in the same position as [3H]CsA mixed with albumin and myoglobin (presumably as free unbound drug). These findings indicate that CsA normally binds to an intraerythrocytic protein similar in molecular size to calf thymus cyclophilin (Mr 15,000). By equilibrium dialysis, the purified erythrocyte proteins calmodulin (Mr 16,700) and cytochrome b5 (Mr 15,000) failed to bind CsA. By equilibrium dialysis, [3H] CsA did bind to column fractions containing the CsA-binding protein, but [125I]CsA did not, suggesting that attachment to CsA occurs at or near a carbon-carbon double bond in an unusual nine-carbon amino acid of CsA. These results have important implications for CsA therapy with regard to distribution space, pharmacokinetics, and a possible protein-receptor mechanism of action.
为研究不同个体血液中环孢素（CsA）的红细胞饱和容量差异现象，对洗涤红细胞的溶血产物进行检测，以确定是否存在CsA结合蛋白。采用凝胶过滤柱色谱法对口服CsA患者的溶血产物进行分析，大部分与红细胞相关的CsA以单一峰形式洗脱，其相对分子质量（Mr）为15,000 - 17,000，与血红蛋白和碳酸酐酶不同。体外向溶血产物中添加的[³H]CsA也以类似方式洗脱。向溶血产物中添加的[¹²⁵I]CsA在与白蛋白和肌红蛋白混合的[³H]CsA相同位置（可能作为游离未结合药物）洗脱得晚得多。这些发现表明，CsA通常与一种分子大小与小牛胸腺亲环蛋白（Mr 15,000）相似的红细胞内蛋白结合。通过平衡透析，纯化的红细胞蛋白钙调蛋白（Mr 16,700）和细胞色素b5（Mr 15,000）未能结合CsA。通过平衡透析，[³H]CsA确实与含有CsA结合蛋白的柱级分结合，但[¹²⁵I]CsA未结合，这表明CsA的结合发生在CsA一种特殊的九碳氨基酸的碳 - 碳双键处或其附近。这些结果对于CsA治疗在分布空间、药代动力学以及可能的蛋白 - 受体作用机制方面具有重要意义。
