Cation binding to SERCA

Sarcoendoplasmic reticulum Ca2+-ATPase (SERCA) is a transmembrane pump that plays an important role in transporting calcium into the sarcoplasmic reticulum (SR). While Ca2+ binds SERCA with micromolar affinity, Mg2+ and K+ compete with Ca2+ binding, yet the molecular bases for these competing ions' influence on SERCA function has not been established. We therefore used three in silico methods to resolve molecular determinants of cation binding in the canonical site I and II Ca2+ binding sites: 1) triplicate molecular dynamic (MD) simulations of Mg2+, Ca2+ and K+-bound SERCA. 2) mean spherical approximation (MSA) theory to determine the affinity and selectivity of cation binding to the MD-resolved structures and 3) state models of SERCA turnover informed from MSA-derived affinity data. Our key findings are that a) coordination at sites I and II are optimized for Ca2+ and to a lesser extent for Mg2+ and K+, as determined by MD-derived cation-amino acid oxygen and bound water configurations, b) the impaired coordination and high desolvation cost for Mg2+ precludes favorable Mg2+ binding relative to Ca2+, while K+ has limited capacity to bind site I, c) Mg2+ most likely acts as inhibitor and K+ as intermediate in SERCA's reaction cycle, based on a state model of SERCA turnover. These findings provide a quantitative basis for SERCA function that leverages molecular-scale thermodynamic data and rationalize enzyme activity across broad range of K+, Ca2+ and Mg2+ concentrations.