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EMBO J. 2013 Dec 11;32(24):3231-43. doi: 10.1038/emboj.2013.250. Epub 2013 Nov 22.

SERCA mutant E309Q binds two Ca(2+) ions but adopts a catalytically incompetent conformation.

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1] Department of Molecular Biology and Genetics, Centre for Membrane Pumps in Cells and Disease - PUMPKIN, Danish National Research Foundation, Aarhus University, Aarhus, Denmark [2] Department of Biomedicine, Aarhus University, Aarhus, Denmark.


The sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) couples ATP hydrolysis to transport of Ca(2+). This directed energy transfer requires cross-talk between the two Ca(2+) sites and the phosphorylation site over 50 Å distance. We have addressed the mechano-structural basis for this intramolecular signal by analysing the structure and the functional properties of SERCA mutant E309Q. Glu(309) contributes to Ca(2+) coordination at site II, and a consensus has been that E309Q only binds Ca(2+) at site I. The crystal structure of E309Q in the presence of Ca(2+) and an ATP analogue, however, reveals two occupied Ca(2+) sites of a non-catalytic Ca2E1 state. Ca(2+) is bound with micromolar affinity by both Ca(2+) sites in E309Q, but without cooperativity. The Ca(2+)-bound mutant does phosphorylate from ATP, but at a very low maximal rate. Phosphorylation depends on the correct positioning of the A-domain, requiring a shift of transmembrane segment M1 into an 'up and kinked position'. This transition is impaired in the E309Q mutant, most likely due to a lack of charge neutralization and altered hydrogen binding capacities at Ca(2+) site II.

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