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Transmembrane Cu(i) P-type ATPase pumps are electrogenic uniporters

Dalton Trans. 2020; 
Nisansala Abeyrathna, Sameera Abeyrathna, M Thomas Morgan, Christoph J Fahrni, Gabriele Meloni
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Mutagenesis Services … Experimental Expression and purification of wild-type CopA and mutants For recombinant expression, the synthetic DNA (Genscript Inc.) encoding codon-optimized EcCopA (Uniprot accession number: Q59385) was cloned into a pET-52b(+) vector with an N- Get A Quote

摘要

Cu(i) P-type ATPases are transmembrane primary active ion pumps that catalyze the extrusion of copper ions across cellular membranes. Their activity is critical in controlling copper levels in all kingdoms of life. Biochemical and structural characterization established the structural framework by which Cu-pumps perform their function. However, the details of the overall mechanism of transport (uniporter vs. cotransporter) and electrogenicity still remain elusive. In this work, we developed a platform to reconstitute the model Cu(i)-pump from E. coli (EcCopA) in artificial lipid bilayer small unilamellar vesicles (SUVs) to quantitatively characterize the metal substrate, putative counter-ions and charge translo... More

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