Estimation of inter-binding-site distances in sarcoplasmic reticulum (Ca2+ + Mg2+)-ATPase under occluded and non-occluded conditions

Abstract

Distances between binding sites in skeletal sarcoplasmic reticulum (Ca2+ + Mg2+)-ATPase were estimated by measuring energy transfer between a luminescent lanthanide probe for the calcium sites (Eu3+) and suitable acceptors. A distance measurement between the two calcium binding sites of the enzyme was made with and without MgATP, corresponding to two different states in the hydrolytic cycle. We found that without ATP, the inter-calcium-site distance is about 0.9 nm, while in the presence of MgATP these sites are 0.05 nm to 0.1 nm closer. We also estimated the distance from the calcium sites to the hydrolytic site using CrATP as an acceptor; this distance is approximately 1.8 nm. These measurements provide the start for a description of intramolecular movement during the transport cycle, and should be of use in mapping out the three-dimensional structure of the Ca2+ pump.