To enhance the mechanical strength of hydroxyapatite (HAP), a nano-HAP/β-sheet fibroin (nano-HAP/SF) composite was synthesized by mixing nano-HAP sols with SF sols. A β-sheet structural SF was prepared by a salt-resolving method. The nano-HAP sols was prepared by a co-precipitation method. The phase composition of HAP and nano-HAP/SF composites were investigated by X-ray diffraction and Fourier transform infrared spectroscopy (FTIR). Their morphologies were observed by scanning electron microscopy. The mechanical strength of the nano-HAP/SF composite was measured by a universal material detector. The bioactivity and biocompatibility of the obtained composites were confirmed by tests in simulated body fluid (SBF) and by a culture of rat osteoblasts cell. Results showed that the obtained SF is a typical silk II with β-sheet conformation. The FTIR spectra analysis indicated that the chemical bonds were formed between SF and HAP in the nano-HAP/SF. The result of SBF test demonstrated that the needle-like crystals of HAP were formed on the surface of the sample and the result of cell culture showed that the rat osteoblasts grown and reproduced well on the surface of the composite. The compression strength of SP/HAP reached 63 MPa.