Identification of an Arg-Leu-Arg tripeptide that contributes to the binding interface between the cytokine MIF and the chemokine receptor CXCR4

Michael Lacy, Christos Kontos, Markus Brandhofer, Kathleen Hille, Sabine Groning, Dzmitry Sinitski, Priscila Bourilhon, Eric Rosenberg, Christine Krammer, Tharshika Thavayogarajah, Georgios Pantouris, Maria Bakou, Christian Weber, Elias Lolis, Juergen Bernhagen*, Aphrodite Kapurniotu*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

27 Citations (Web of Science)

Abstract

MIF is a chemokine-like cytokine that plays a role in the pathogenesis of inflammatory and cardiovascular disorders. It binds to the chemokine-receptors CXCR2/CXCR4 to trigger atherogenic leukocyte migration albeit lacking canonical chemokine structures. We recently characterized an N-like-loop and the Pro-2-residue of MIF as critical molecular determinants of the CXCR4/MIF binding-site and identified allosteric agonism as a mechanism that distinguishes CXCR4-binding to MIF from that to the cognate ligand CXCL12. By using peptide spot-array technology, site-directed mutagenesis, structure-activity-relationships, and molecular docking, we identified the Arg-Leu-Arg (RLR) sequence-region 87-89 that-in three-dimensional space-'extends' the N-like-loop to control site-1-binding to CXCR4. Contrary to wildtype MIF, mutant R87A-L88A-R89A-MIF fails to bind to the N-terminal of CXCR4 and the contribution of RLR to the MIF/CXCR4-interaction is underpinned by an ablation of MIF/CXCR4-specific signaling and reduction in CXCR4-dependent chemotactic leukocyte migration of the RLR-mutant of MIF. Alanine-scanning, functional competition by RLR-containing peptides, and molecular docking indicate that the RLR residues directly participate in contacts between MIF and CXCR4 and highlight the importance of charge-interactions at this interface. Identification of the RLR region adds important structural information to the MIF/CXCR4 binding-site that distinguishes this interface from CXCR4/CXCL12 and will help to design MIF-specific drug-targeting approaches.
Original languageEnglish
Article number5171
Number of pages17
JournalScientific Reports
Volume8
DOIs
Publication statusPublished - 26 Mar 2018

Keywords

  • MIGRATION INHIBITORY FACTOR
  • SECONDARY STRUCTURE ANALYSES
  • FAST INTERACTION REFINEMENT
  • CRYSTAL-STRUCTURE
  • OLIGOMERIZATION STATE
  • NMR CHARACTERIZATION
  • INTERNATIONAL UNION
  • MOLECULAR DOCKING
  • CELL RECRUITMENT
  • STRUCTURAL BASIS

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