Structural role of countertransport revealed in Ca 2+ pump crystal structure in the absence of Ca 2+

Ca 2+ -ATPase of sarcoplasmic reticulum is an ATP-powered Ca 2+ pump but also a H + pump in the opposite direction with no demonstrated functional role. Here, we report a 2.4-Å-resolution crystal structure of the Ca 2+ -ATPase in the absence of Ca 2+ stabilized by two inhibitors, dibutyldihydroxybenzene, which bridges two transmembrane helices, and thapsigargin, also bound in the membrane region. Now visualized are water and several phospholipid molecules, one of which occupies a cleft between two transmembrane helices. Atomic models of the Ca 2+ binding sites with explicit hydrogens derived by continuum electrostatic calculations show how water and protons fill the space and compensate charge imbalance created by Ca 2+ -release. They suggest that H + countertransport is a consequence of a requirement for maintaining structural integrity of the empty Ca 2+ -binding sites. For this reason, cation countertransport is probably mandatory for all P-type ATPases and possibly accompanies transport of water as well.