The kinetics of dissociation of the inhibitor of nucleoside transport, nitrobenzylthioinosine, from the high-affinity binding sites of cultured hamster cells.
The kinetics of dissociation of the inhibitor of nucleoside transport, nitrobenzylthioinosine, from the high-affinity binding sites of cultured hamster cells.
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核苷转运抑制剂硝基苄基硫代肌苷从培养仓鼠细胞的高亲和力结合位点解离的动力学。
DOI：
10.1042/bj2160299
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
A. Paterson
中科院分区：
文献类型：
--
作者：
R. Koren;C. Cass;A. Paterson
硝基苄基硫代肌苷 (NBMPR) 与质膜上一组高亲和力位点的结合可抑制各种类型动物细胞中的核苷转运。这项工作检测了 [3H]NBMPR 与培养的 Nil 8 仓鼠成纤维细胞和源自 Nil 8 的病毒转化克隆 (Nil SV) 细胞的核苷转运蛋白的结合。用完整的 Nil 8 和 Nil SV 细胞以及膜制剂进行的实验表明，这两个细胞系在结合位点的细胞含量方面存在显着差异，并且这些位点对 NBMPR 的亲和力仅有轻微差异。 Nil 8 和 Nil SV 细胞每个细胞分别拥有 (4.2-8.0) X 10(5) 和 (2.0-4.0) X 10(6) 个位点，而用完整细胞和膜制备物获得的位点结合 NBMPR 解离常数相似，范围为 0.29 至 1.5 nM。 Dilazep 是一种与 NBMPR 结构无关的核苷转运的有效抑制剂，在平衡条件下测试时，它似乎与 NBMPR 竞争与高亲和力位点的结合，抑制 NBMPR 与 Nil 8 和 Nil SV 细胞结合的 Ki 值分别为 15 +/- 4 和 32 +/- 4 nM。 NBMPR 从 Nil SV 膜制剂的结合位点 - NBMPR 复合物上解离是一级衰减过程，速率常数为 0.68 +/- 0.26 min-1。 NBMPR 从膜制剂和完整细胞的结合位点复合物中解离的速率在地拉西普存在下显着降低，而在渗透性尿苷存在下增加。这些结果表明，在平衡条件下地拉西普获得的表观竞争抑制动力学不应被解释为地拉西普与 NBMPR 相同的位点结合，而是两种抑制剂与核苷转运蛋白上密切相关的位点的结合。类似地，尿苷似乎也结合到与 NBMPR 结合位点不同的位点。
Nucleoside transport in various types of animal cells is inhibited by the binding of nitrobenzylthioinosine (NBMPR) to a set of high-affinity sites on the plasma membrane. This work examined the binding of [3H]NBMPR to the nucleoside transporters of cultured Nil 8 hamster fibroblasts and of cells of a virus-transformed clone (Nil SV) derived from Nil 8. Experiments conducted with intact Nil 8 and Nil SV cells and with membrane preparations indicated that the two lines differed significantly in the cellular content of binding sites and only slightly in the affinities of these sites for NBMPR. Nil 8 and Nil SV cells possessed (4.2-8.0) X 10(5) and (2.0-4.0) X 10(6) sites per cell respectively, whereas the dissociation constants of site-bound NBMPR obtained with intact cells and with membrane preparations were similar, ranging from 0.29 to 1.5 nM. Dilazep, a potent inhibitor of nucleoside transport that is structurally unrelated to NBMPR, appeared to compete with NBMPR for binding to the high-affinity sites when tested under equilibrium conditions with Ki values for inhibition of NBMPR binding to Nil 8 and Nil SV cells respectively of 15 +/- 4 and 32 +/- 4 nM. The dissociation of NBMPR from the binding site--NBMPR complex of Nil SV membrane preparations was a first-order decay process with a rate constant of 0.68 +/- 0.26 min-1. The rate of dissociation of NBMPR from the binding-site complex of membrane preparations and intact cells was decreased significantly in the presence of dilazep and increased in the presence of the permeant uridine. These results suggest that the apparent competitive-inhibition kinetics obtained for dilazep under equilibrium conditions should not be interpreted as binding of dilazep to the same site as NBMPR but rather as binding of the two inhibitors to closely associated sites on the nucleoside transporter. Similarly, uridine also appears to bind to a site separate from the NBMPR-binding site.