Modeling a dehalogenase fold into the 8-angstrom density map for Ca2+-ATPase defines a new domain structure

Members of the large family of P-type pumps use active transport to maintai n gradients of a wide variety of cations across cellular membranes. Recent structures of two P-type pumps at 8-Angstrom resolution have revealed the a rrangement of transmembrane helices but were insufficient to reveal the arc hitecture of the cytoplasmic domains. However, recent proposals of a struct ural homology with a superfamily of hydrolases offer a new basis for modeli ng these domains. In the current work, we have extended the sequence compar ison for the superfamily and delineated domains in the 8-Angstrom density m ap of Ca2+-ATPase. The homology suggests a new domain structure for Ca2+-AT Pase and, specifically, that the phosphorylation domain adopts a Rossman fo ld. Accordingly, the atomic structure of L-2 haloacid dehalogenase has been fitted into the relevant domain of Ca2+-ATPase. The resulting model sugges ts the existence of two ATP sites at the interface between two domains. Bas ed on this new model, we are able to reconcile numerous results of mutagene sis and chemical cross-linking within the catalytic domains. Furthermore, w e have used the model to predict the configuration of MS ATP at its binding site. Based on this prediction, we propose a mechanism,involving a change in Mg2+ liganding, for initiating the domain movements that couple sites of ion transport to ATP hydrolysis.