A knottin scaffold directs the CXC-chemokine-binding specificity of tick evasins

J Biol Chem. 2019 Jul 19;294(29):11199-11212. doi: 10.1074/jbc.RA119.008817. Epub 2019 Jun 5.


Tick evasins (EVAs) bind either CC- or CXC-chemokines by a poorly understood promiscuous or "one-to-many" mechanism to neutralize inflammation. Because EVAs potently inhibit inflammation in many preclinical models, highlighting their potential as biological therapeutics for inflammatory diseases, we sought to further unravel the CXC-chemokine-EVA interactions. Using yeast surface display, we identified and characterized 27 novel CXC-chemokine-binding evasins homologous to EVA3 and defined two functional classes. The first, which included EVA3, exclusively bound ELR+ CXC-chemokines, whereas the second class bound both ELR+ and ELR- CXC-chemokines, in several cases including CXC-motif chemokine ligand 10 (CXCL10) but, surprisingly, not CXCL8. The X-ray crystal structure of EVA3 at a resolution of 1.79 Å revealed a single antiparallel β-sheet with six conserved cysteine residues forming a disulfide-bonded knottin scaffold that creates a contiguous solvent-accessible surface. Swapping analyses identified distinct knottin scaffold segments necessary for different CXC-chemokine-binding activities, implying that differential ligand positioning, at least in part, plays a role in promiscuous binding. Swapping segments also transferred chemokine-binding activity, resulting in a hybrid EVA with dual CXCL10- and CXCL8-binding activities. The solvent-accessible surfaces of the knottin scaffold segments have distinctive shape and charge, which we suggest drives chemokine-binding specificity. These studies provide structural and mechanistic insight into how CXC-chemokine-binding tick EVAs achieve class specificity but also engage in promiscuous binding.

Keywords: chemokine; chemotaxis; crystal structure; evasin; host-pathogen interaction; immune response; inflammation; knottin; protein-protein interaction; tick.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chemokines, CXC / metabolism*
  • Crystallography, X-Ray
  • Cystine-Knot Miniproteins / metabolism*
  • Protein Binding
  • Protein Conformation
  • Receptors, Chemokine / genetics
  • Receptors, Chemokine / isolation & purification
  • Receptors, Chemokine / metabolism*
  • Species Specificity
  • Ticks / classification
  • Ticks / metabolism*
  • Yeasts / genetics


  • Chemokines, CXC
  • Cystine-Knot Miniproteins
  • Receptors, Chemokine

Associated data

  • PDB/6I31