Nucleoside transport in mammalian cell membranes - III. Kinetic and chemical modification studies of cytosine-arabinoside and uridine transport in hamster cells in culture

Transport of the nucleoside analog cytosine-arabinoside (CAR) in transformed hamster cells in culture has been studied in conditions of minimal metabolic conversion. Uptake (zero-trans in) properties at 20°C over a limited range of CAR concentrations were characterized by a K_m of 350 μm and a maximal velocity (V) of 780 μm·min^-1 (V/K_m=2.28 min^-1). Equilibrium exchange at 20°C over a wider range of concentrations was best described by a saturable component with a K_m of 500 μm and a v of 1230 μm·min^-1 (V/K_m=2.26 min^-1) and either a saturable component of high K_m or a nonsaturable component of k=0.3 min^-1. For the saturable component, the v/K_m values were similar in both procedures. CAR transport was inhibited by various metabolizable nucleosides. Uptake of some of these nucleosides was inhibited by CAR. CAR transport and uridine uptake were inhibited in a reversible but partially competitive fashion by high affinity probes like S-(p-nitrobenzyl-6-mercaptoinosine (NBMI) (K_i<0.5 nm) and in an irreversible fashion by SH reagents such as N-ethylmaleiimide (NEM). The organomercurial p-hydroxymercuribenzene sulfonate (pMBS) markedly stimulated transport of these nucleosides, but also markedly potentiated the inhibitory effects of either NBMI or NEM. These effects are interpreted either in terms of models which invoke allosteric properties or in terms of two transport systems which display distinct chemical susceptibilities to externally added probes.