Reconciling disparate views of template-directed nucleation through measurement of calcite nucleation kinetics and binding energies

Abstract

The physical basis for how macromolecules regulate the onset of mineral formation in calcifying tissues is not well established. A popular conceptual model assumes the organic matrix provides a stereochemical match during cooperative organization of solute ions. In contrast, another uses simple binding assays to identify good promoters of nucleation. Here, we reconcile these two views and provide a mechanistic explanation for template-directed nucleation by correlating heterogeneous nucleation barriers with crystal-substrate-binding free energies. We first measure the kinetics of calcite nucleation onto model substrates that present different functional group chemistries (carboxyl, thiol, phosphate, and hydroxyl) and conformations (C11 and C16 chain lengths). We find rates are substrate-specific and obey predictions of classical nucleation theory at supersaturations that extend above the solubility of amorphous calcium carbonate. Analysis of the kinetic data shows the thermodynamic barrier to nucleation is reduced by minimizing the interfacial free energy of the system, γ. We then use dynamic force spectroscopy to independently measure calcitesubstrate-binding free energies, ΔG b . Moreover, we show that within the classical theory of nucleation, γ and ΔG b should be linearly related. The results bear out this prediction and demonstrate that low-energy barriers to nucleation correlate with strong crystal-substrate binding. This relationship is general to all functional group chemistries and conformations. These findings provide a physical model that reconciles the long-standing concept of templated nucleation through stereochemical matching with the conventional wisdom that good binders are good nucleators. The alternative perspectives become internally consistent when viewed through the lens of crystal-substrate binding.B iological systems are unique in their ability to organize minerals into functional materials with complex patterns and architectures. A substantial body of evidence suggests specialized macromolecules, particularly proteins (1, 2) and carbohydrates (3, 4), provide preferential sites for nucleation to direct the placement, timing, and orientation of crystals (5), both intra-and extracellular. Within the biomineralization community, the conventional view of biologically directed nucleation is that macromolecular matrices present an interfacial match to the crystal lattice that assists in forming the crystal nucleus. This cooperative view of directed nucleation is rooted in the collective action of multiple residues that guide the organization of ions into a configuration defining the energetic minimum for the system. A series of in vitro observations have reinforced this picture by showing that highly ordered organic monolayers can control the location and orientation of calcite crystals precipitated from solution (6). In this approach, good templates are revealed through a direct functional assay, i.e., nucleation. Over the years, this view of mineralization, both in the context of natural stru...

Energetic basis for the molecular-scale organization of bone

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“…S5). In addition, one is hard pressed to explain why the clear stereochemical matching of the matrix to the mineral plays no role in directing growth given the demonstration of this effect in even simple systems (44,46). Thus, both mechanisms may work in concert to exert the observed control.…”

Amorphous Phase Mediated Crystallization: Fundamentals of Biomineralization

“…So, reducing the interfacial energy or increasing the supersaturation would greatly promote the nucleation rate, which has been widely corroborated in many crystallization systems. With respect to biomineralization systems, the biomatrix and NMPs are believed to facilitate crystal nucleation by reducing the interfacial energy of nucleation [182,183] or increasing the local supersaturation by charge attractions [70,[184][185][186][187].…”

Probing the energetics of organic–nanoparticle interactions of ethanol on calcite

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