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

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

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Radcliffe Department of Medicine Division of Cardiovascular Medicine, University of Oxford, Oxford OX3 7BN, United Kingdom.
Serono Pharmaceutical Research Institute, 1228 Geneva, Switzerland.
Department of Chemistry, University of Oxford, Oxford OX1 3TA, United Kingdom.
Serono Pharmaceutical Research Institute, 1228 Geneva, Switzerland
Radcliffe Department of Medicine Division of Cardiovascular Medicine, University of Oxford, Oxford OX3 7BN, United Kingdom


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.


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

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