Ion binding and selectivity of the rotor ring of the Na+-transporting V-ATPase.

Abstract

The vacuole-type ATPases (V-ATPases) are proton pumps in various intracellular compartments of eukaryotic cells. Prokaryotic V-ATPase of Enterococcus hirae, closely related to the eukaryotic enzymes, provides a unique opportunity to study ion translocation by V-ATPases because it transports Na(+) ions, which are easier to detect by x-ray crystallography and radioisotope experiments. The purified rotor ring (K-ring) of the E. hirae V-ATPase binds one Na(+) ion per K-monomer with high affinity, which is competitively inhibited by Li(+) or H(+), suggesting that the K-ring can also bind these ions. This finding is also supported by the K-ring structure at 2.8 A in the presence of Li(+). Association and dissociation rates of the Na(+) to and from the purified K-ring were extremely slow compared with the Na(+) translocation rate estimated from the enzymatic activity, strongly suggesting that interaction with the stator subunit (I-subunit) is essential for Na(+) binding to /release from the K-ring.