Characterization of the effects of phosphorylation by CK2 on the structure and binding properties of human HP1β

•HP1β contains two primary sites for CK2 with distinct phosphorylation kinetics.•Localized conformational and dynamic changes occur at the sites of phosphorylation.•HP1β phosphorylated at S89 and S175 retains its ability to bind chromatin.

Proteins of the Heterochromatin Protein 1 (HP1) family are regulators of chromatin structure and genome function in eukaryotes. Post-translational modifications expand the repertoire of the chemical diversity of HP1 proteins and regulate their activity. Here, we investigated the effect of phosphorylation by Casein kinase 2 (CK2) on the structure, dynamics and binding activity of human HP1β. We show that Ser89 in the hinge region is the most effective substrate, followed by Ser175 at the C-terminal tail. Phosphorylation at these sites results in localized conformational changes in HP1β that do not compromise the ability of the protein to bind chromatin.