Determining the molecular drivers of species-specific interferon-stimulated gene product 15 interactions with nairovirus ovarian tumor domain proteases
(A) Overlay of the KUPEV, GANV, and CCHFV OTUs. The OTU “selectivity helix” is indicated by a red oval, with closeup views of the helix in each structure shown to the right. (B) Impact of single or multiple mutations within the helix on the activity of the KUPEV, GANV, and CCHFV OTUs towards Ub-AMC and human ISG15-AMC. Values represent the mean ± standard deviation of two independent experiments. (C) Cleavage assays of OTU mutants with human and sheep proISG15. Samples from each timepoint were run on BioRad Mini-PROTEAN® TGX™ gels and visualized by Coomassie staining. (D) Western blot analysis of OTU activity on conjugated human or sheep ISG15. Cell lysates containing conjugated human or sheep ISG15 was incubated with purified OTUs, and deISGylase activity was assessed by Western blot.