Modification of erythrocyte Na+/Li+ countertransport kinetics by two types of thiol group

Erythrocyte Na+/Li+ countertransport activity is decreased by reagents that react with thiol groups. An understanding of the role of these groups in control of Na+/Li+ countertransport may help to explain its association with disease states. The effect of thiol reactive agents on the kinetic parameters of Na+/Li+ countertransport has not previously been described. In choline medium, N-ethylmaleimide (NEM) and iodoacetamide (IAamide) cause a rapid decrease of about 40% in Km for external sodium (Km(So)) that is complete in 10 s with a much smaller change in Vmax and an increase in the Vmax/Km ratio. In Na medium, NEM and IAamide both cause a rapid decrease in Km(So) and Vmax. With NEM the partial reduction in Vmax is complete in 100s although the NEM is sufficient to reduce Vmax up to 15 min. With IAamide the decrease in Vmax is initially slower but it continues apparently towards complete inhibition. These results indicate at least two types of thiol group controlling Na+/Li+ countertransport kinetics. The type 1 thiol reaction is Na independent and causes an increase in the apparent rate constant for Na association with the unloaded carrier so that Vmax/Km rises and Km(So) decreases. The type 2 thiol reaction is facilitated by Na at the outside ion-binding site and causes a decrease in Vmax, possibly by total blockage of carriers with IAamide but by a different mechanism with NEM such as reduced turnover rate.