ATP dependent charge movement in ATP7B Cu+-ATPase is demonstrated by pre-steady state electrical measurements.
ATP7B is a copper dependent P-type ATPase, required for copper homeostasis. Taking advantage of high yield heterologous expression of recombinant protein, we investigated charge transfer in ATP7B. We detected charge displacement within a single catalytic cycle upon ATP addition and formation of phosphoenzyme intermediate. We attribute this charge displacement to movement of bound copper within ATP7B. Based on specific mutations, we demonstrate that enzyme activation by copper requires occupancy of a site in the N-terminus extension which is not present in other transport ATPases, as well as of a transmembrane site corresponding to the cation binding site of other ATPases.
Department of Chemistry Ugo Schiff, University of Florence, 50019 Sesto Fiorentino, Italy. francesco.tadini@unifi.it
Francesco
F
Tadini-Buoninsegni
Gianluca
G
Bartolommei
Maria Rosa
MR
Moncelli
Rajendra
R
Pilankatta
David
D
Lewis
Giuseppe
G
Inesi
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000327:Adsorption; D000818:Animals; D019556:COS Cells; D027682:Cation Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D000073840:Copper-Transporting ATPases; D004553:Electric Conductivity; D004560:Electricity; D004579:Electron Transport; D006801:Humans; D008670:Metals / Q000378:metabolism; D017434:Protein Structure, Tertiary
D027682:Cation Transport Proteins; D008670:Metals; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C000612464:ATP7B protein, human; D000073840:Copper-Transporting ATPases