11.2. Ion pumping[[Student version, January 17, 2003]] 421
11010050 100 150 200 250 300outward sodium flux, arbitrary unitstime, minutesion pumps
shut downFigure 11.5:(Experimental data.) Flux of sodium ions out of a cuttlefish axon after electrical stimulation. At
the beginning of the experiment the axon was loaded with radioactive sodium, then placed in ordinary sea water;
the loss of radioactivity was then monitored. During the interval represented by the arrow, the axon was exposed to
the toxin dinitrophenol (DNP), temporarily shutting down sodium pumping. Later the toxin was washedawaywith
fresh seawater, and ion pumping spontaneously resumed. The horizontal axis gives the time after end of electrical
stimulation; the vertical scale gives the rate at which radioactively labeled sodium left the axon. [Data from Hodgkin
&Keynes, 1955.]
passive, Ohmic part of the fluxes unchanged. Moreover, even with the cell’s metabolism shut down,
pumping resumes when one injects the cellular energy-storing molecule ATP into the cell.
Tosummarize, the results described above pointed to a hypothesis:
Aspecific molecular machine embedded in cell membranes hydrolyzes ATP,
then uses some of the resulting free energy to pump sodium ions out of the cell.
Atthe same time the pump imports potassium, partially offsetting the loss of
electric charge from the exported sodium.
(11.11)
The pump operates only when sodium and ATP are available on its inner side and potassium is
available on its outer side. If any of these are cut off, the cell slowly reverts to the ion concentrations
appropriate for equilibrium.
Idea 11.11 amounts to a remarkably detailed portrait of the membrane pump, considering that
in 1955 no specific membrane constituent was even known to be a candidate for this job. Clearly
somethingwaspumping those ions, but there are thousands of transmembrane proteins in a living
cell membrane, and it was hard to find for the right one. But in 1957, the physiologist J. Skou
isolated a single membrane protein with ATPase activity from crab leg neurons. Carefully control-
ling the ion content of his solutions, Skou found that to hydrolyze ATP, his enzyme required both
sodium and potassium, the same behavior Hodgkin, Katz, and Ussing had found for whole nerve
axons (Figure 11.6). Skou concluded that his enzyme must have separate binding sites for both
sodium and potassium. For this and other reasons, he correctly guessed that it was the missing
sodium pump.
Additional experiments confirmed Skou’s hypotheses: Remarkably, it is possible to prepare a
pure lipid bilayer, introduce the purified pump protein, the necessary ions, and ATP, then watch as