Biochemistry, Department of

 

Document Type

Article

Date of this Version

2023

Citation

Chem. Sci., 2023, 14, 6059-6078.

DOI: 10.1039/d3sc00347g

Comments

Open access. © 2023 The Author(s). Published by the Royal Society of Chemistry

Abstract

Transmembrane P-type ATPase pumps catalyze the extrusion of transition metal ions across cellular lipid membranes to maintain essential cellular metal homeostasis and detoxify toxic metals. Zn()-pumps of the P-type subclass, in addition to Zn , select diverse metals (Pb, Cdand Hg) at their transmembrane binding site and feature promiscuous metal-dependent ATP hydrolysis in the presence of these metals. Yet, a comprehensive understanding of the transport of these metals, their relative translocation rates, and transport mechanism remain elusive. We developed a platform for the characterization of primary-active Zn()-pumps in proteoliposomes to study metal selectivity, translocation events and transport mechanism in real-time, employing a “multi-probe” approach with fluorescent sensors responsive to diverse stimuli (metals, pH and membrane potential). Together with atomic-resolution investigation of cargo selection by X-ray absorption spectroscopy (XAS), we demonstrate that Zn()-pumps are electrogenic uniporters that preserve the transport mechanism with 1-, 2- and 3-row transition metal substrates. Promiscuous coordination plasticity, guarantees diverse, yet defined, cargo selectivity coupled to their translocation.

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