Structural and functional insights into a Zn2+-transporting P-type ATPase.

Zinc is essential for life yet toxic in excess, rendering homeostatic control vital for all organisms. Transition metal transporting P-type (PIB-) ATPase proteins are critical for this balance but remain poorly understood at the molecular level. Here, we present the inward-open structure of a dimeric zinc-transporting PIB-type ATPase, resolved at 3.6 Å resolution. Biochemical analyses and molecular dynamics simulations suggest that the N-terminal tail, harboring two so-called metal binding domains (MBDs), auto-inhibits the ATPase core and serves as a metal-sensitive sensor, releasing inhibition when intracellular metal levels rise. Contrary to the prevailing uptake models, the structure reveals an electronegatively charged uptake region that directly exposes a metal-binding cysteine of the invariant CPC-motif. We propose that a histidine-rich segment, positioned after the MBD closest to the core, donates ions to the CPC site. Consequently, an intricate interplay between the regulatory and ion-providing N-terminus and the ATPase core is present, safeguarding against zinc toxicity. Collectively, our findings shed light on the regulation and transport mechanism of PIB-ATPases, with implications for fundamental biology and the potential to inform future biotechnological and translational applications.