Identification and functional analysis of three novel osteogenic peptides isolated from tilapia scale collagen hydrolysate

On the basis of our previous study that 133 peptides were identified from the tilapia scale peptide-calcium chelate, the potential osteogenic peptide monomers through the calcium-binding properties of peptides and molecular docking were screened, and the osteogenic activity and active mechanism of the peptides were further researched in this study. Three highly osteogenic peptides GPAGPHGPVG (844.4191 Da), APDPFRMY (995.4534 Da), and TPERYY (827.3813 Da) were screened. Molecular docking showed that the three osteogenic peptides had the same interaction sites in the epidermal growth factor receptor (EGFR), namely ARG 285, GLN 8, GLY 317, THR 406, and HIS 409. Compared to the blank control group, within 50 μg/mL of GPAGPHGPVG, APDPFRMY, and TPERYY increased the proliferation of MC3T3-E1 cells by changing the cell proportion in the S and G2/M phases, and the alkaline phosphatase (ALP) activity of MC3T3-E1 cells treated with 50 μg/mL of the three active peptides increased by 25%, 37%, and 56%, respectively. The three active peptides at 10 μg/mL concentration significantly promoted the mineralization of osteoblasts, and the mineralized calcium nodules increased by 166%, 161%, and 111%, respectively. TPERYY significantly increased the expression of osteogenic genes (osteocalcin (OCN), type I collagen (Col I α) and transcription factor (OSX)). Moreover, TPERYY significantly increased the mRNA and protein expression of β-catenin to 1.39 and 2.6 times of the blank control group, respectively, while decreased the mRNA and protein expression of glycogen synthesis kinase (GSK3β) to 1.6 and 2.3 times of the blank control group, respectively. This study provides a theoretical basis for using GPAGPHGPVG, APDPFRMY, and TPERYY peptides as functional foods to prevent osteoporosis.