Emu Eggshell Proteins DCA-1 and DCA-2 Shape Calcite Growth, Study Finds

Researchers have mapped how two proteins from the emu eggshell help control the chemical environment in which calcium carbonate crystals form, offering new structural and functional evidence on biomineralization in a living relative of some of the most ancient bird lineages.

The study, published in The Science of Nature, focused on Dromaius novaehollandiae, the emu, a paleognath bird whose lineage diverged early from other birds. The team combined structural, chemical, proteomic and computational approaches to characterize the eggshell and identify the intramineral proteins that regulate crystal growth within its mineral phase and organic matrix.

Using scanning electron microscopy with energy-dispersive X-ray spectroscopy, Fourier-transform infrared spectroscopy and X-ray diffraction, the researchers showed that calcite is the predominant mineral phase in the emu eggshell. However, they identified structural and chemical differences compared with the chicken, Gallus gallus, particularly in the organization of the internal fibrillar network and in components associated with the organic matrix.

Proteomic analysis then pinpointed biomineralization-related proteins, including DCA-1 and DCA-2, as candidates for regulating the mineralizing environment. To test whether these proteins actually interact with the ionic building blocks of the shell, the team carried out microtitration and crystal growth assays in systems containing calcium ions and carbonate or bicarbonate species.

Those assays showed that the protein fractions exhibit functional activity in the Ca²⁺ and CO₃²⁻/HCO₃⁻ systems, suggesting differential affinities for calcium and carbonate species. In other words, the two proteins do not simply bind the same ions in the same way; their behavior points to distinct roles in managing the local chemistry during nucleation and growth of calcite.

To visualize how this might happen at the molecular level, the researchers used AlphaFold3 to produce the first three-dimensional structural models of DCA-1 and DCA-2 interacting with Ca²⁺ and CO₃²⁻. The models revealed distinct interaction patterns: DCA-1 displayed a more distributed stabilization network for carbonate, whereas DCA-2 showed more concentrated regions associated with calcium and a more localized interaction with carbonate ions.

Taken together, the authors argue, the results suggest that DCA-1 and DCA-2 participate complementarily in regulating the mineralizing microenvironment during shell formation made of calcite. One protein appears oriented toward stabilizing carbonate species across broader structural regions, while the other concentrates on calcium-associated interactions, a division of labor that could help fine-tune crystal nucleation and growth as the shell forms.

Because the emu belongs to the paleognaths, an early-diverging bird group, the findings contribute to understanding ancestral mechanisms of biomineralization and add new structural and functional evidence on intramineral proteins beyond the well-studied chicken eggshell. The work is based on laboratory characterization, proteomics, in vitro assays and computational modeling, and the functional conclusions rest on the observed activities of protein fractions and on modeled interaction patterns rather than direct observation inside a forming shell.

Subject of Research: Proteins DCA-1 and DCA-2 in emu eggshells and their roles in regulating calcium carbonate biomineralization

Article Title: DCA-1 and DCA-2 proteins of Dromaius novaehollandiae are involved in regulating the chemical microenvironment during the nucleation and growth of CaCO₃ in eggshell formation

Article References: Guardado-Urbina, A. G., Loredo-Portales, R., Rosales-Hoz, M. J., Moreno, A., & Cuéllar-Cruz, M. (2026). DCA-1 and DCA-2 proteins of Dromaius novaehollandiae are involved in regulating the chemical microenvironment during the nucleation and growth of CaCO₃ in eggshell formation. The Science of Nature, 113(6), Article 141. https://doi.org/10.1007/s00114-026-02188-y

Image Credits: AI Generated

DOI: 10.1007/s00114-026-02188-y

Keywords: emu, eggshell, biomineralization, calcite, DCA-1, DCA-2, calcium carbonate, proteomics, AlphaFold3, paleognaths, crystal growth, organic matrix