Autoimmune diseases impose a significant global health burden, and current anti-inflammatory therapies, such as corticosteroids, are associated with adverse effects that limit their long-term use. Consequently, there is a need for novel anti-inflammatory therapeutics. Poxviruses produce cytokine-binding proteins that modulate the host immune response and represent a relatively unexplored source of anti-inflammatory agents. One such protein, M-T7 from Myxoma virus, is a homolog of the rabbit interferon-gamma (IFN-γ) receptor and binds to IFN-γ as well as a range of mammalian chemokines. M-T7 is the only known poxviral immunomodulatory protein reported to exhibit bivalent sequestration of both interferons and chemokines, two families of cytokines critical to inflammatory signalling. It has shown therapeutic promise in mouse models of organ transplantation and wound healing, and is therefore a candidate for clinical translation. However, challenges in crystallising M-T7 have thus far prevented structural characterisation of its cytokine-binding mechanism.
Recombinant M-T7 was expressed in HEK293-6E cell culture and purified by immobilised metal-ion affinity chromatography and size-exclusion chromatography. M-T7 was successfully crystallised, and diffraction data were collected on the MX1 beamline at the Australian Synchrotron. The unligated structure of M-T7 was solved to 2.5 Å by X-ray crystallography. Small-angle X-ray scattering (SAXS) data are consistent with M-T7 forming a tetrameric assembly in solution, similar to other interferon-binding proteins. Surface plasmon resonance (SPR) was used to characterise interactions between M-T7 and a panel of human chemokines. SPR binding analyses suggested low-micromolar affinities for select chemokines, although some previously reported interactions were not reproduced.
These findings provide the first high-resolution structural insights into M-T7 and indicate its oligomeric state in solution. Together with binding kinetic data, this work advances understanding of the structure–function relationships underlying M-T7-mediated cytokine sequestration and immunomodulation, supporting its continued investigation as a potential therapeutic for autoimmune and inflammatory diseases.