The Landscape of Osteocalcin\nProteoforms Reveals Distinct\nStructural and Functional Roles of Its Carboxylation Sites

Human osteocalcin (OC) undergoes reversible, vitamin\nK-dependent\nγ-carboxylation at three glutamic acid residues, modulating\nits release from bones and its hormonal roles. A complete understanding\nof OC roles and structure–activity relationships is still lacking,\nas only uncarboxylated and few differently carboxylated variants have\nbeen considered so far. To fill this lack of knowledge, a comprehensive\nexperimental and computational investigation of the structural properties\nand calcium-binding activity of all the OC variants is reported here.\nSuch a comparative study indicates that the carboxylation sites are\nnot equivalent and differently affect the OC structure and interaction\nwith calcium, properties that are relevant for the modulation of OC\nfunctions. This study also discloses cooperative effects and provides\nstructural and mechanistic interpretation. The disclosed peculiar\nfeatures of each carboxylated proteoform strongly suggest that considering\nall eight possible OC variants in future studies may help rationalize\nsome of the conflicting hypotheses observed in the literature.