Long-Timescale Molecular Dynamics Reveal a Coordination-Biased Conformational Selection Mechanism for Sorcin...

Sorcin is a dimeric penta-EF-hand Ca²⁺-binding protein that regulates intracellular Ca²⁺ homeostasis through Ca²⁺-dependent conformational activation and target recognition, and it has also been implicated in multidrug resistance in cancer. Although crystal structures have defined the apo inactive and Ca²⁺-bound active states of Sorcin, the transition pathways connecting these states and the conformational ensembles populated under each condition remain poorly understood. Here, we used long-timescale allatom molecular dynamics simulations on Anton 3, totaling ~90 μs, to define the Ca²⁺-coupled conformational landscape of dimeric human Sorcin at atomic resolution. Starting from the Ca²⁺-bound structure, we directly observed the transition from the active to the inactive state following Ca²⁺ removal, demonstrating that loss of Ca²⁺ coordination is sufficient to drive inactivation on the microsecond timescale. Simulations initiated from the Ca²⁺-bound crystal structure with retained ions unexpectedly revealed ultrafast Ca²⁺ dissociation and rebinding at all EF-hand sites, indicating weak intrinsic Ca²⁺ affinity and highly dynamic ion exchange. In complementary simulations initiated from the apo structure, Sorcin spontaneously sampled active-like conformations even in the absence of stable Ca²⁺ binding, supporting a conformational selection mechanism in which Ca²⁺ shifts the population toward pre-existing active states rather than inducing the transition de novo. Across all conditions, we also observed pronounced and persistent structural asymmetry between the two protomers, revealing that the Sorcin homodimer is dynamically heterogeneous despite its symmetric crystal structures. Together, these results support a coordination-biased conformational selection model for Sorcin activation, in which weak and rapidly exchanging Ca²⁺ binding stabilizes, rather than induces, the active state. This work provides a dynamic framework for understanding Sorcin function as a fast Ca²⁺ sensor and offers broader mechanistic insight into activation principles of EF-hand Ca²⁺-binding proteins.