Contribution of the Hydroxyl Group and Phenol Ring of Tyrosine 780 of the Alpha-Subunit to NA+ Binding by the Na/K Pump

Abstract

By applying well designed oscillating electrical pulse to a single frog skeletal muscle with voltage-clamp and double vaseline gap techniques, we obtained pump current on both positive and negative half cycle. The frequency of the oscillating pulses remains at 50 Hz which is comparable with the physiological turnover rate of the Na/K pump. Different magnitudes of voltage (from 30mV to 65mV with a 5mV step) were applied to the muscle. The results showed that the pump current as well as the charge mediated by pump moved across the plasma cell membrane were voltage dependent and reached a saturation status when the pulse was greater than 55 mV. In this situation, the total amount of pump-mediated charge remained roughly the same even with the changing magnitude of the activation pulses. The charge on the positive half cycle and negative half cycle which represent the Na extrusion and K intrusion respectively, has a ratio less than but close to 3:2. After adding ouabain, the currents were recorded each 5 minutes. The results showed that the total pump-mediated charge decreased monotonically and reached minimum which was around 0 after 20 minutes. This indicated that this oscillating electrical pulse induced current was highly sensitive to and can be totally eliminated by ouabain, a specific Na/K pump blocker.