ructural Basis for Differential Binding of the Interleukin-8 Monomer and Dimer to the CXCR1 N-Domain: Role of Coupled Interactions and Dynamics

摘要

ABSTRACT: Interleukin-8 (IL-8 or CXCL8) plays a critical role in orchestrating the immune response bynbinding and activating the receptor CXCR1 that belongs to the GPCR class. IL-8 exists as both monomersnand dimers, and both bind CXCR1 butwith differential affinities. It iswell established that themonomer is thenhigh-affinity ligand and that the interactions between the ligandN-loop and receptorN-domain play a criticalnrole in determining binding affinity. In order to characterize the structural basis of differential binding of thenIL-8 monomer and dimer to the CXCR1 N-domain, we analyzed binding-induced NMR chemical shift andnpeak intensity changes and show that they are exquisitely sensitive and can provide detailed insights into thenbinding process.We used three IL-8 variants, a designedmonomer, a trapped disulfide-linked dimer, andWTnat dimeric concentrations. NMR data for the monomer show that nonsequential residues that span the entirenN-loop are involved in the binding process and that the binding is mediated by a network of extensive directnand indirect coupled interactions. Interestingly, in the case of WT, binding induces dissociation of thendimer-receptor complex to the monomer-receptor complex, and in the case of the trapped dimer, bindingnresults in increased global conformational flexibility. Increased dynamics is evidence of unfavorableninteractions, indicating that binding of the WT dimer triggers conformational changes that disruptndimer-interface interactions, resulting in its dissociation. These results together provide evidence thatnbinding is not a localized event but results in extensive coupled interactions within the monomer and acrossnthe dimer interface and that these interactions play a fundamental role in determining binding affinity.