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Adenine transport and binding in cultured mammalian cells deficient in adenine phosphoribosyltransferase.

ABSTRACT

Rapid kinetic techniques were employed to measure the transport of adenine in adenine phosphoribosyltransferase-deficient L929 and Chinese hamster ovary (CHO) cells in zero-trans entry and exit and equilibrium exchange procedures. The kinetic parameters of transport were computed by fitting appropriate integrated rate equations to time courses of transmembrane equilibration of radiolabeled adenine. Adenine transport conformed to the simple carrier model with directional symmetry and equal mobility of loaded and empty carrier. The Michaelis-Menten constants and maximum velocities for various strains of L929 cells fell between 2.3 and 3.5 mM and 90 and 150 pmol/microliters of cell water per s, respectively, values similar to those previously reported for CHO and Novikoff hepatoma cells. The corresponding values for hypoxanthine transport in L929 cells were 413 microM and 16 pmol/microliters of cell water per s. Adenine transport velocities were directly proportional to adenine concentrations between 0.03 and 50 microM in both CHO and Novikoff cells. The results indicate that adenine is transported in these cells by a single, low-affinity, high-capacity transporter. Adenine transport was inhibited by hypoxanthine in some cell strains, but not in others. Adenine also rapidly bound to L929 cells in a saturable manner (KD = 18 microM), presumably to the cell surface (about 3 X 10(7) sites per cell).