For controlled bone regeneration, two-dimensional materials are essential because they provide an isolated area for bone regeneration and protect the surrounding tissues. Excellent mechanical and hydrophilic qualities have made graphene oxide (GO) a common choice for bone regeneration. A biomimetic mineralization process combined with one-step vacuum filtering created mechanically strong and biocompatible two-dimensional (2D) membranes. Silk fibroin (SF) and hydroxyapatite (HA) increased the membranes' mechanical strength and hy-drophilicity on the surface. Osteoblast adhesion, differentiation, and mineralization improved with little HA administered. A calvarial defect model was used to test the impact of the produced composite membranes on bone formation in vivo after they were implanted. Histological examination demonstrates that bone regeneration may be accelerated by GO/SF/HA composite membranes without needing endogenous cytokines. GO/SF/HA nanocomposite membranes with distinctive porous architectures, exceptional mechanical characteristics, and excellent bone regeneration capability are viable nanocomposites for bone regeneration.