Reversible homodimerization of a small leucine-rich proteoglycan PRELP modulates the interactions with TGFβ1 and IGFI-R.

Small leucine-rich proteoglycans (SLRPs) are responsible for various biological functions via their multiple interactions with various proteins. Although some SLRPs form homodimers, it remains unclear how homodimerization affects the molecular functions of SLRPs. Herein, we report the molecular basis and functional significance of homodimerization of proline/arginine-rich end leucine-rich repeat protein (PRELP), which is a member of SLRP family. We used biophysical techniques to validate the premise that PRELP forms a reversible homodimer in a concentration-dependent manner. Because the spatial aggregation propensity calculation indicated that the concave surface of the leucine-rich repeat domain of PRELP is highly hydrophobic, we speculated that this hydrophobicity predominantly drives PRELP homodimerization. To address this hypothesis, we replaced several amino acid residues that showed high spatial aggregation propensity values with charged residues identified by the Rosetta energy calculation, which resulted in disruption of PRELP homodimerization. Molecular dynamics simulation further verified the molecular basis of the monomerizing mutation as well as an unexpected dimerizing mutation. To demonstrate the influence of reversible homodimerization on PRELP interactions, we conducted interaction analysis using the designed PRELP mutants. We found that the monomerizing mutation of PRELP enhanced interactions with transforming growth factor β1 and insulin-like growth factor I receptor, suggesting that the functions of PRELP that rely on these interactions are moderated via reversible homodimerization.