STRUCTURAL, DYNAMIC, AND ELECTRONIC INVESTIGATION OF THE BIOMINERALIZATION OF CALCIUM CARBONATE AGGREGATES IN A PROTEIN MATRIX

In this study, acidic protein matrices were investigated for their potential role in regulating calcium carbonate biomineralization through modulation of hydration within and around the protein. How hydration and ion association are coupled during early aggregation remains incompletely understood. Atomistic molecular dynamics simulations were used to examine zebrafish Starmaker (STM), a protein involved in otolith formation, in explicit water containing Ca²⁺ and CO₃²⁻. Protein dynamics, hydration, ion distributions, and clustering were characterized using RMSD, RMSF, radius of gyration, radial distribution functions (RDFs; g(r)), spatial distribution functions (SDFs), and interaction analyses. Water showed distinct organization in the protein interior, at the external interface, and around the ions, while aspartate (ASP) residues generated spatially heterogeneous hydration patterns. Ion distribution analyses indicated that Ca²⁺ and CO₃²⁻ preferentially occupied distinct regions around ASP residues rather than forming a single co-localized layer. Relative to apo STM, the ion-containing system showed a broader RMSD distribution without evidence of simple uniform compaction from the radius of gyration. Ion aggregation was limited but compositionally organized: smaller clusters were carbonate-rich and became progressively enriched in Ca²⁺ as they grew. This trend was consistent with Ca²⁺–CO₃²⁻ attraction, although quantitative energy estimates require further validation. Small CaCO₃-rich aggregates, consistent with early putative nuclei, were observed within the protein interior. Together, these results suggest that ASP-rich regions reorganize local hydration and spatially partition the ions, while STM conformational redistribution creates dynamic environments that promote ion recruitment and aggregate growth. These putative nuclei may grow through further aggregation toward larger amorphous or crystalline domains.