The Isolated C-Terminal Domain of Ring1B Is a Dimer Made of Stable, Well-Structured Monomers†

The Ring1B is a core subunit protein of the PRC1 (polycomb repressive complex 1), which plays key roles in the regulation of the Homeobox gene expression, X-chromosome inactivation, stem cell self-renewal, and tumorigenesis. The C-terminal region of Ring1B interacts with RYBP, a transcriptional repressor in transiently transfected cells, and also with M33, another transcriptional repressor involved in mesoderm patterning. In this work, we show that the C-terminal domain of Ring1B, C-Ring1B, is a dimer in solution, with a dissociation constant of 200 μM, as shown by NMR, ITC, and analytical gel filtration. Each monomer is stable at physiological conditions in a wide pH range (∼5 kcal mol -1 at 298 K), with a well-formed core and a spherical shape. The dimer has a high content of α-helix and β-sheet, as indicated by FTIR spectra, and it is formed by the mutual docking of the preformed folded monomers. Since the C-terminal region is important for interaction with other proteins of the PRC1, the dimerization and the presence of those well-structured monomers might be a form of regulation.