Following Transient Conformational Adaptations during Peptide-Guided Calcium Phosphate Biomineralization with Hyperpolarized NMR.

Peptide-guided biomineralization is a key process involved in the formation of calcium phosphate (CaP) in bone, yet the peptide-ion encounter complexes at the onset of this process have remained structurally invisible: methods with sufficient resolution to decipher these intermediates would already have lost them to precipitation. Here, we resolve how a mineralization peptide adapts its conformation to recruit inorganic ions during the earliest stages of CaP formation. Using a peptide derived from the calcium-binding domain of secreted phosphoprotein 1 (SPP1), a key regulator of bone formation, we find that Ca2+ association reorganizes the conformational ensemble toward states with increased solvent exposure of Ca2+-coated Asp-rich stretches. These polyionic microenvironments subsequently exhibit spectral perturbations upon phosphate recruitment, consistent with the formation of encounter complexes during the onset of CaP nucleation at the peptide-solvent interface. Access to these transient states is enabled by NMR sensitized with hyperpolarized water (HyperW), which provides a two-order-of-magnitude sensitivity gain and reduces multidimensional acquisition times from hours to secondsfast enough to capture peptide-ion assemblies immediately after encounter and before precipitation. Together, these results reveal a stepwise mechanism by which an intrinsically disordered mineralization peptide senses and organizes calcium and phosphate ions at the onset of biomineralization.