Plagemann P G, Aran J M, Wohlhueter R M, Woffendin C
Department of Microbiology, University of Minnesota, Minneapolis 55455.
Biochim Biophys Acta. 1990 Feb 16;1022(1):103-9. doi: 10.1016/0005-2736(90)90405-d.
Time courses of transmembrane equilibration of 2-chloroadenosine, 2'-deoxyadenosine, 3'-deoxyadenosine, cytidine and 2'-deoxycytidine were measured by rapid kinetic techniques in human erythrocytes under equilibrium exchange and zero-trans conditions. The kinetic parameters for transport were computed by fitting appropriate integrated rate equations to the data pooled for seven concentrations and compared to the kinetic parameters for uridine, adenosine, thymidine and formycin B transport determined previously for human erythrocytes under comparable experimental conditions. The transport of all nucleosides conformed to the simple carrier model and was directionally symmetric. The Michaelis-Menten constants for equilibrium exchange (Kee) ranged from 22 microM for 2-chloroadenosine to about 4 mM for cytidine and the maximum velocities (Vee) differed in a similar manner, so that the first-order rate constants (Vee/Kee) were similar for all nucleosides. The kinetic parameters for 2'-deoxyadenosine transport were similar to those for adenosine transport, whereas the lack of the 3'-OH group greatly reduced the affinity of 3'-deoxyadenosine (cordycepin) for the carrier. 2', 3'-Dideoxynucleosides were transported less than 1% as efficiently as 2'- and 3'-deoxynucleosides. Thus, the 2'- and 3'-OH groups play an important role in nucleoside transport. The mobility of the carrier when loaded with pyrimidine nucleosides (reflected by Vee) was 5-10-times greater than that of the empty carrier, whereas the mobility of the adenosine-loaded or 2'-deoxyadenosine-loaded carrier was about equal to that of the empty carrier. Loading the carrier with 2-chloroadenosine or 3'-deoxyadenosine actually decreased its mobility. Thus, the differential mobility of the loaded and empty carrier differs greatly with the nucleoside substrate. The mobility of the loaded carrier as well as Kee increased with a decrease in lipid solubility of the nucleoside substrate, but the relationship was complex.
在平衡交换和零转运条件下，采用快速动力学技术测定了人红细胞中2-氯腺苷、2'-脱氧腺苷、3'-脱氧腺苷、胞苷和2'-脱氧胞苷的跨膜平衡时程。通过将适当的积分速率方程拟合到七种浓度的数据集中，计算出转运的动力学参数，并与先前在可比实验条件下测定的人红细胞中尿苷、腺苷、胸苷和间型霉素B转运的动力学参数进行比较。所有核苷的转运均符合简单载体模型，且具有方向对称性。平衡交换的米氏常数（Kee）范围从2-氯腺苷的22微摩尔到胞苷的约4毫摩尔，最大速度（Vee）也以类似方式不同，因此所有核苷的一级速率常数（Vee/Kee）相似。2'-脱氧腺苷转运的动力学参数与腺苷转运的相似，而3'-OH基团的缺失大大降低了3'-脱氧腺苷（虫草素）对载体的亲和力。2',3'-双脱氧核苷的转运效率不到2'-和3'-脱氧核苷的1%。因此，2'-和3'-OH基团在核苷转运中起重要作用。装载嘧啶核苷时载体的迁移率（由Vee反映）比空载载体大5-10倍，而装载腺苷或2'-脱氧腺苷的载体的迁移率与空载载体大致相等。用2-氯腺苷或3'-脱氧腺苷装载载体实际上会降低其迁移率。因此，装载和空载载体的差异迁移率因核苷底物而异。装载载体的迁移率以及Kee随核苷底物脂溶性的降低而增加，但这种关系很复杂。
