Novel Calcium-Binding Peptide from Bovine Bone Collagen Hydrolysates and Its Potential Pro-Osteogenic Activity via Calcium-Sensing Receptor (CaSR)

Scope: This paper aims to explore the osteogenic activity and potential mechanism of the peptide-calcium chelate, and provides a theoretical basis for peptide-calcium chelates as functional foods to prevent or improve osteoporosis.

Methods and results: In this research, a novel peptide (Phe-Gly-Leu, FGL) with a high calcium-binding capacity is screened from bovine bone collagen hydrolysates (CPs), calcium binding sites of which mainly included carbonyl, amino and carboxyl groups. The FGL-Ca significantly enhances the osteogenic activity of MC3T3-E1 cells (survival rate, differentiation, and mineralization). The results of calcium fluorescence labeling and molecular docking show that FGL-Ca may activate calcium-sensing receptor (CaSR), leading to an increase in intracellular calcium concentration, then enhancing osteogenic activity of MC3T3-E1 cells. Further research found that FGL-Ca significantly promotes the mRNA and protein expression levels of CaSR, transforming growth factor β (TGF-β1), TGF-β-type II receptor (TβRII), SMAD2, SMAD3, osteocalcin (OCN), Alkaline Phosphatase (ALP), osteoprotegrin (OPG), and collagen type I (COLI). Subsequently, in the signal pathway intervention experiment, the expression levels of genes and proteins related to the TGF-β1/SMAD2/3 signaling pathway that are promoted by FGL-Ca are found to decrease.

Conclusions: These results suggest that FGL-Ca may activate CaSR, increase intracellular calcium concentration, and activate TGF-β1/SMAD2/3 signaling pathway, which may be one of the potential mechanisms for enhancing osteogenic activity.

MC3T3-E1 cells; calcium-binding capacity; calcium-binding peptide; calcium-sensing receptor; collagen hydrolysates; osteogenic activity.