Structure and Function of Copper Transport ATPases

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Metal micronutrients, Fe, Cu, Zn, etc., play fundamental catalytic and structural functions in proteins. Proper metal levels are achieved by the coordinate action of specific chaperones and transmembrane transporters. The goal of this project is to understand the fundamental biochemistry involved in transmembrane heavy metal transport by ubiquitous P1B-type ATPases. Alkali metals (Na+, K+, Ca2+) are free (hydrated) in the cellular milieus. As such, they reversibly interact with and are transported by channels, co-transporters, and pumps. In contrast, heavy metals are bound to chaperones/chelating molecules with high association constants. This project aims to understand how heavy metals are delivered to their transporters, how they are coordinated with affinities likely higher than that of their chaperones, and how they are subsequently released from the transporter. In these studies, Archaeoglobus fulgidus CopA will be used as a model Cu+-ATPase. Experiments will be performed to test whether CopZ, the A. fulgidus Cu+ chaperone, interacts with CopA delivering Cu+ to transmembrane metal binding sites. The stoichiometry of metal binding to transmembrane sites will be established along with the atomic coordination and the arrangement of coordinating amino acid side chains. The project will also examine the structural mechanism of allosteric regulation by metal binding to cytoplasmic domains. Finally, the fourth goal of the project is to establish the complete atomic resolution structure of a Cu+ transporting ATPase. This would greatly enhance understanding the mechanism of heavy metal transport. Broader Impact: The project will contribute to understanding the principles governing metal binding, translocation and release mechanisms of metal transporters. The research will contribute to ongoing educational activities including undergraduate teaching, curriculum development and summer research. Graduate and undergraduate students form the core of the laboratory and will perform major parts of this project. This work will also impact a close collaboration with K-12 programs at our institution. This ranges from guiding high school students working in the lab to participation in high school teachers training.