Session H51: Biomaterials: Structure, Function, Design III

Sea Urchin Biomineralization – Formation of Intricate Single Calcite Crystals via Amorphous Precursors

Biomineralization of sea urchin skeletal elements results in complex structures with smooth, curved surfaces that diffract as single calcite crystals 1. This is achieved by a crystallization pathway involving amorphous precursor phases 2,3. Similar processes are now recognized in various other organisms. Yet, the dynamics of mineral rearrangement and the energetic landscape of this transformation are still poorly understood. We addressed these questions studying the crystallization of biogenic and synthetic ACCs by state-of-the-art calorimetric, spectroscopic and scattering methods. The skeletal elements of the sea urchin Paracentrotus lividus are composed of ACC, organics, a small amount of water and calcite 4 . Insight into the interplay between these components is gained by HR-XRPD of skeletal elements annealed at elevated temperatures. Complementarily, by mapping the distribution of ACC . H2O, ACC and calcite in growing spines by X-PEEM we demonstrated variable transformation kinetics across the spine. In-vitro experiments show that the effect of water, as well as organic and inorganic additives, on the stability of synthetic ACC is primarily kinetic [5]. A key finding with relevant to
biomineralization is that although water drastically changes the crystallization enthalpy, the overall ACC thermodynamic stability is independent of the hydration level due to enthalpy-entropy compensation. The rate of the transformation, on the other hand, is critically dependent on the water content; water increases ion mobility, resulting in higher kinetic instability [5].

1. Berman a, et al. (1993) Science 259, 776–9
2. Politi Y, Arad T, Klein E, Weiner S, Addadi L (2004) Science 306, 1161–4
3. Politi Y, et al. (2008) Proc Natl Acad Sci U S A 105, 17362–6
4. Albéric M, et al. (2018) Cryst Growth Des 18, 2189–201
5. Albéric M, et al. (2017) Adv Sci 1701000. doi:10.1002/advs.201701000
6. Jensen ACS, et al. (2018) J Phys Chem C 122, 3591–8
7. Zou Z, et al. (2018) J Mater Chem B 6, 449–57   

Multistep Crystallization Pathways for Protein Crystals and Colloidal Assemblies

While much is now known about the extraordinary complexity and structural diversity possible for crystals of proteins, nanoparticles and colloids, far less is known about the process by which these crystals form. Simple "classical" models of nucleation and growth may hold for the self-assembly of simple crystal structures, but what of crystals with large unit cells? We present recent results from computer simulation demonstrating that crystallization pathways in protein solutions or entropic colloidal fluids may be as diverse and complex as the resulting crystals, with crystallization following a remarkable variety of multistep pathways that include fluid-fluid transitions. We explore the role of shape, entropy, and, in the case of proteins, both specific and nonspecific interactions in selecting the pathway.   

Design of Functional Protein Membranes

Protein surfaces are composed of sub-nanometer domains, with different degrees of hydrophilicity. These domains play a crucial role in protein assembly, ligand recognition, and drug docking. We explored here the surface domains to disperse enzymes in organic solvents where their activity can be enhanced using random heteropolymers that mimic unstructured proteins in membraneless organelles. The analysis of the correlations between polar domains with the polar components of amphiphilic random heteropolymers mediated by water elucidates the formation of protein-polymer complexes with a core (protein)-shell(polymer) morphology that help stabilize the proteins’ structures and preserve their enzymatic activities in unfavorable solvents as observed in recent experiments (1). At a more coarse-grained resolution, we find that the proteins selectively favor the binding of random copolymers with similar monomer sequences (2). The balance between the energetic and entropy gains in polymer adsorption is determined by the spatial distribution of the polar and non-polar domains and the average composition of the polymers.

(1) B. Panganiban, et al. Science 359: 1239-1243 (2018)
(2) T. Nguyen, et al. PNAS 26, 6578-6583 (2018)   

The Growth Mechanisms and Biomimetics of Tooth Enamel

A very fundamental part of biomineralization is the complex extracellular macromolecular framework in which mineralization occurs, such as the collagen fibrils in bone and dentin, polysaccharides in nacre and amelogenin and non-amelogenin proteins in dental enamel. Unlike other mineralized tissues, such as bone and dentin, mature enamel is acellular and cannot regenerate itself after substantial mineral loss. Biomimetic enamel regrowth is a significant topic in material science and dentistry as an alternative approach for the treatment of defects in dental enamel. We have developed protocols for superficial biomimetic enamel regrowth, based on a novel amelogenin-chitosan hydrogel. Amelogenin is a critical protein for controlling the organized growth of apatite crystals in enamel. We expanded upon the concept of biomineralization to design smaller amelogenin-inspired peptides with conserved functional domains for clinical translation. The synthetic peptides displayed a characteristic nanostructured scaffold reminiscent of ‘nanospheres’ seen in the enamel matrix and effectively controlled apatite nucleation in vitro. Following application of the peptides to sectioned human molar teeth, a robust, oriented, synthetic aprismatic enamel was observed in situ. There was a two-fold increase in the hardness and modulus of the regrown enamel-like apatite layers and an increase in the attachment of the tooth-regrown layer interface compared to control samples. Repeated peptide applications generated multiple enamel-like HAP layers of limited thickness produced by epitaxial growth in which c-axis oriented nanorods evolved on the surface of native enamel. We report that peptide analogues with active domains can effectively regulate the orientation of regenerated HAP layers to influence functional response.   

Engineering materials inspired by nature

Magnetic field aligned freeze casting is a novel method to fabricate porous, anisotropic ceramic scaffolds with a hierarchy of architectural alignment in multiple directions. This concept was inspired by the structure of trabecular bone and the spiraling nature of the narwhal tusk. A weak rotating magnetic field applied normal to the ice growth direction in a uniaxial freezing apparatus allowed the manipulation of magnetic nanoparticles to create different pore structures and channels with long-range order in directions parallel and perpendicular to the freezing direction. Porous scaffolds consisting of different host ceramics as particles or platelets (hydroxyapatite (HA), ZrO2, Al2O3, or TiO2) mixed with varying concentrations of Fe3O4 nanoparticles were fabricated by freeze casting under no magnetic field, a static or rotating magnetic field. In the magnetic field direction, the compressive strength and stiffness of the scaffolds containing was doubled.