The Mechanism Of Cholesterol Binding Sequence Mediating The Interaction Of MCRP With Diverse Ligands

C-reactive protein(CRP) is a risk marker of inflammatory diseases and a mediator in inflammatory foci. Accumulating evidence has demonstrated that the function of CRP depends on its conformation and localization. mCRP appears to be the major conformation of CRP that regulates local inflammation. Our previous studies have shown that a single sequence motif in mCRP,i.e.cholesterol binding sequence(CBS; a.a. 35-47), is responsible for mediating its interactions with diverse ligands. However, little is known about the mechanism of CBS-mediated interactions between mCRP and its ligands. Here, we expressed and purified mCRP mutants in E. coli. We found that a complete deletion of CBS significantly decreased the binding capacity of mCRP with ligands. Molecular dynamics(MD) simulation result showed that CBS existed as an intrinsic disordered region, which provided a structural explanation for the CBS-mediated interactions. Furthermore, by stimulating endothelial cells with mCRP and observing the monocyte adhesion, we found that the presence of CBS peptide could significantly inhibit the effects of mCRP on activation of endothelial cells. Meanwhile, intraperitoneal co-injection mCRP and CBS peptide tomice model markedly inhibited mCRP-induced IL-6 release. These results demonstrated that CBS peptide was able to inhibit the proinflammatory actions of mCRP both in vitro and in vivo. CBS can be identified as the major recognition site of mCRP. We suggest that thismotif may be a target to adjust the proinflammatory actions ofmCRP and that optimized CBS peptide can be developed as a potential inhibitor of mCRP.