Use of membrane vesicles to estimate the numbers of system y⁺ and system L amino acid transporters in human erythrocytes

We have used equilibrium values for L-leucine and L-lysine uptake by right-side-out vesicles to estimate the membrane abundance (sites/cell) of Na(+)-dependent amino acid transport systems L and y+ in human erythrocytes. All of the intravesicular space was accessible to L-leucine, as judged by comparisons with uridine uptake via the equilibrative nucleoside transporter (10(4) sites/cell). In contrast, only 28% of the total intravesicular space was accessible to L-lysine uptake via system y+. Since human erythrocyte membranes generate an average of approximately 1000 vesicles/cell, these data provide evidence that system L is a relatively high-abundance membrane transport protein in human erythrocytes, while system y+ is present in smaller amounts (approximately 300 copies/cell). Calculated turnover numbers for L-lysine transport by system y+ at 37 degrees C are 24 s-1 for zero-trans influx and 150 s-1 for equilibrium-exchange influx.