Enzyme-assisted mineralization of calcium phosphate: exploring confinement for the design of highly crystalline nano-objects.

In hard tissues of vertebrates, calcium phosphate (CaP) biomineralization is a fascinating process that combines specific physicochemical and biochemical reactions, resulting in the formation of extracellular matrices with elegant nanoarchitectures. Although several “biomimetic” strategies have been developed for the design of mineralized nanostructured biointerfaces, the control of the crystallization process remains intricate. Herein, we report an innovative approach to overcome this challenge by generating, in situ, CaP precursors in confined medium. For this purpose, we explore a combination of (i) the layer-by-layer assembly, (ii) the template-based method and (iii) the heterogenous enzymatic catalysis. We show the possibility to embed active alkaline phosphatase in a nanostructured multilayered film and to induce the nucleation and growth of CaP compounds in different conditions. Importantly, we demonstrate that the degree of confinement of active enzymes modulate the crystal phase: from spheroid-shaped amorphous CaP to crystalline platelet-shaped hydroxyapatite. This leads to the synthesis of highly anisotropic mineralized nanostructures, mechanically stable and with controlled dimensions, composition and crystal phase. The present study provides a straightforward, yet powerful, way to design anisotropic nanostructured materials, including slef-supported framework, which may be used in broad biomedical applications.