Biophysical characterization of the CXC chemokine receptor 2 ligands.

The chemokines of the immune system act as first responders by operating as chemoattractants, directing immune cells to specific locations of inflamed tissues. This promiscuous network is comprised of 50 ligands and 18 receptors where the ligands may interact with the receptors in various oligomeric states i.e., monomers, homodimers, and heterodimers. Chemokine receptors are G-protein coupled receptors (GPCRs) present in the membrane of immune cells. The migration of immune cells occurs in response to a concentration gradient of the ligands. Chemotaxis of neutrophils is directed by CXC-ligand (CXCL) activation of the membrane bound CXC chemokine receptor 2 (CXCR2). CXCR2 plays an important role in human health and is linked to disorders such as autoimmune disorders, inflammation, and cancer. Yet, despite their important role, little is known about the biophysical characteristics controlling ligand:ligand and ligand:receptor interaction essential for biological activity. In this work, we study the homodimers of three of the CXCR2 cognate ligands, CXCL1, CXCL5, and CXCL8. The ligands share high structural integrity but a low sequence identity. We show that the sequence diversity has evolved different binding affinities and stabilities for the CXC-ligands resulting in diverse agonist/antagonist behavior. Furthermore, CXC-ligands fold through a three-state mechanism, populating a folded monomeric state before associating into an active dimer.

A Small Contribution to a Large System: The Leptin Receptor Complex.

Obesity is a classified epidemic, increasing the risk of secondary diseases such as diabetes, inflammation, cardiovascular disease, and cancer. The pleiotropic hormone leptin is the proposed link for the gut-brain axis controlling nutritional status and energy expenditure. Research into leptin signaling provides great promise toward discovering therapeutics for obesity and its related diseases targeting leptin and its cognate leptin receptor (LEP-R). The molecular basis underlying the human leptin receptor complex assembly remains obscure, due to the lack of structural information regarding the biologically active complex. In this work, we investigate the proposed receptor binding sites in human leptin utilizing designed antagonist proteins combined with AlphaFold predictions. Our results show that binding site I has a more intricate role in the active signaling complex than previously described. We hypothesize that the hydrophobic patch in this region engages a third receptor forming a higher-order complex, or a new LEP-R binding site inducing allosteric rearrangement.