Ca²⁺-induced self-assembly of Bombyx mori silk sericin into a nanofibrous network-like protein matrix for directing controlled nucleation of hydroxylapatite nano-needles

Bone biomineralization is a well-regulated protein-mediated process where hydroxylapatite (HAP) crystals are nucleated with preferred orientation within self-assembled protein matrix. Mimicking this process is a promising approach to the production of bone-like protein/mineral nanocomposites for bone repair and regeneration. Towards the goal of fabricating such nanocomposites from sericin, a protein spun by <i>Bombyx mori</i> (<i>B.mori</i>) silkworm, and bone mineral HAP, for the first time we investigated the chemical mechanism underpinning the synergistic processes of the conformational change/self-assembly of <i>B.mori</i> sericin ( <b><i>BS</i></b> ) as well as the nucleation of HAP on the resultant self-assembled <b><i>BS</i></b> matrix. We found that <b><i>BS</i></b> , rich in anionic amino acid residues, could bind Ca²⁺ ions from the HAP precursor solution through electrostatic attraction. The Ca²⁺binding drove the conformational change of <b><i>BS</i></b> from random coils into β-sheets and its concomitant self-assembly into interconnected nanofibrous network-like protein matrix, which initiated the nucleation and growth of HAP crystals. HAP crystals directed by the resultant self-assembled <b><i>BS</i></b> matrix grew preferentially along their crystallographic c-axis, leading to the formation of HAP nano-needles. The HAP nano-needles in the self-assembled <b><i>BS</i></b> matrix were subsequently aggregated into globules, probably driven by the hydrogen bonding between C=O groups of <b><i>BS</i></b> and O-H groups of HAP nano-needles. The present work sheds light on the chemical mechanisms of <b><i>BS</i></b> self-assembly and the controlled mineralization directed by the self-assembled matrix. We also found that the resultant nanocomposites could promote the osteogenic differentiation of human bone marrow-derived mesenchymal stem cells. Thus our work also generates a biomimetic approach to bone-like silk protein/mineral nanocomposite scaffolds that can find potential applications in bone repair and regeneration.