Plagemann P G, Woffendin C
Department of Microbiology, Medical School, University of Minnesota, Minneapolis 55455.
Biochim Biophys Acta. 1989 Jan 17;1010(1):7-15. doi: 10.1016/0167-4889(89)90177-8.
Formycin B, a C-nucleoside analog of inosine, is not catabolized by human erythrocytes and mouse P388 leukemia cells and is only very inefficiently phosphorylated in these cells. This relative inertness allows the measurement of its transport into and out of the cells uncomplicated by metabolic conversions. We have measured the zero-trans and equilibrium exchange flux of formycin B in these cells by rapid kinetic techniques. The Michaelis-Menten constants and maximum velocities for formycin B transport in both types of cell were similar to those previously reported for uridine and thymidine. Nevertheless, the differential mobility of the substrate-loaded and empty carrier of human erythrocytes was less for formycin B than uridine as substrate. Formycin B influx was inhibited by other nucleosides in accordance with their affinities for the carrier, but unaffected by purines. The inhibition of formycin B influx by nitrobenzylthioinosine and dipyridamole was also identical to that observed with uridine as substrate (IC50 = 10 and 30 nM, respectively). Formycin B accumulated in both types of cell to 30-40% higher concentrations than were present in the medium. This concentrative accumulation was not due to active transport, metabolism or partitioning into membrane lipids. It seems to reflect binding of formycin B to intracellular components, but does not interfere significantly with measurements of its transport.
间型霉素B是肌苷的C - 核苷类似物，它不会被人类红细胞和小鼠P388白血病细胞分解代谢，并且在这些细胞中磷酸化效率极低。这种相对惰性使得在不涉及代谢转化的情况下就能测量其进出细胞的转运情况。我们已通过快速动力学技术测量了间型霉素B在这些细胞中的零转运和平衡交换通量。间型霉素B在这两种细胞中的米氏常数和最大转运速度与先前报道的尿苷和胸苷相似。然而，以间型霉素B为底物时，人类红细胞中底物负载型和空载载体的差异迁移率比以尿苷为底物时要小。间型霉素B的内流受到其他核苷的抑制，抑制程度与其对载体的亲和力一致，但不受嘌呤影响。硝基苄硫肌苷和双嘧达莫对间型霉素B内流的抑制作用也与以尿苷为底物时观察到的相同（IC50分别为10和30 nM）。间型霉素B在这两种细胞中的积累浓度比培养基中的高30 - 40%。这种浓缩积累并非由于主动转运、代谢或分配到膜脂中。它似乎反映了间型霉素B与细胞内成分的结合，但对其转运测量没有显著干扰。
