Conformational Transitions and Alternating-Access Mechanism in the Sarcoplasmic Reticulum Calcium Pump

•All-atom computational pathways of three main conformational transitions in SERCA•Phosphoryl transfer reaction cannot take place without the binding of two calcium ions.•The transition responsible for binding and occlusion of calcium ions is cooperative.•Luminal opening is driven by the cytoplasmic domains and is a two-step process.•Closing of the luminal gate drives dephosphorylation in the catalytic site.•The results elucidate the coupling between ATPhydrolysis and calcium transport.

Ion pumps are integral membrane proteins responsible for transporting ions against concentration gradients across biological membranes. Sarco/endoplasmic reticulum Ca2 +-ATPase (SERCA), a member of the P-type ATPases family, transports two calcium ions per hydrolyzed ATP molecule via an “alternating-access” mechanism. High-resolution crystallographic structures provide invaluable insight on the structural mechanism of the ion pumping process. However, to understand the molecular details of how ATP hydrolysis is coupled to calcium transport, it is necessary to gain knowledge about the conformational transition pathways connecting the crystallographically resolved conformations. Large-scale transitions in SERCA occur at time-scales beyond the current reach of unbiased molecular dynamics simulations. Here, we overcome this challenge by employing the string method, which represents a transition pathway as a chain of states linking two conformational endpoints. Using a multiscale methodology, we have determined all-atom transition pathways for three main conformational transitions responsible for the alternating-access mechanism. The present pathways provide a clear chronology and ordering of the key events underlying the active transport of calcium ions by SERCA. Important conclusions are that the conformational transition that leads to occlusion with bound ATP and calcium is highly concerted and cooperative, the phosphorylation of Asp351 causes a reorganization of the cytoplasmic domains that subsequently drives the opening of the luminal gate, and the reclosing of luminal gate induces a shift in the cytoplasmic domains that subsequently enables the dephosphorylation of Asp351-P. Formation of transient residue–residue contacts along the conformational transitions predicted by the computations provide an experimental route to test the general validity of the computational pathways.

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