Journal:Acta Cryst D:S2059798318015322

Structure of ISG15 from the bat species Myotis davidii and the impact of interdomain ISG15 interactions on viral protein engagement

Caroline Langley, Octavia Goodwin, John V. Dzimianski, Courtney M. Daczkowski and Scott D. Pegan ^[1]

Molecular Tour
Ubiquitin-like interferon-stimulated gene product 15 (ISG15) is a key modulator of interferon responses involved in the innate immune system. This 15 kDa protein is comprised of two ubiquitin (Ub)-like β-grasp three-dimensional folds. Post-translational modifications of viral proteins introduced by ISG15 have been directly observed to affect the function of numerous viral proteins. While ubiquitin is essentially identical across all animals, comparison of ISG15 across species reveals they are relatively divergent, with sequence identity dropping as low as ~58% among mammals. These ISG15 species-species differences have also long been shown to impact viral deISGylase function.

Recently, the structure of the first non-human ISG15 originating from mouse suggested that human structures of ISG15s may not be reflective of other species. Here, the structure of ISG15 from the bat species Myotis davidii solved to 1.37 Å is reported (6mdh). Comparison of this ISG15 structure with those of human and mouse not only underscores the structural impact of ISG15 species-species differences, but also highlights a conserved hydrophobic motif formed between the two domains of ISG15. Using papain-like deISGylase from the severe acute respiratory coronavirus as a probe, the biochemical importance of this interface and its species-species variances on ISG15-protein engagements was illuminated.

. Structures of hISG15 (green; 1z2m^[2])), mISG15 (magenta; 5chf^[3]), and bISG15 (blue 6mdh) are superimposed using the least squared fit of residues comprising the C-terminal domain on each respective protein. .

Apart from the surface differences between bISG15 and those of other structurally solved ISG15, noticeable tertiary differences between these ISG15 structures were observed. When the ISG15s from bat, mouse and human sources are overlaid using a least squared fit of residues comprising the C-terminal domain of each structure, a noticeable difference in the twist about the C-terminal and N-terminal domains becomes apparent. Closer investigation into the interface between the domains of these three proteins reveals the potential molecular drivers for these differences.

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As with previous ISG15 structures, there is an apparent hydrophobic interaction between the N- and C-terminal domains of bISG15. The C-terminal side of the interaction is facilitated by Leu81, Leu99, and Pro143 with the N-terminal side principally using Phe40 supported in its position by Pro38. With the exception of Pro38 that is a histidine residue in hISG15, all these positions are conserved between bISG15 and those of ISG15 originating from mice and humans. However, just as there was a noticeable difference between the hinge regions of mISG15 and hISG15, there is one for bISG15. Specifically, bISG15’s hinge region possesses a type I reverse turn that separates bISG15 from mISG15 and hISG15. Unlike the hinge region of other ISG15s, reverse turn directionally alters the chain’s direction facilitating bISG15 domain-domain orientation observed. . This SCDR primary sequence motif is NCSE in mice and KCDE in humans. With mISG15 and hISG15 lacking the serine hydroxyl group – aspartic carboxylic acid interaction, mISG15 and hISG15 are not able to replicate this tight turn that is present in bISG15.

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However, other mammal ISG15s, such as that of the white tooth shrew, do have the biochemical ability to form the turn. This suggests that this motif is not just solely restricted to bat ISG15s, but could be present in other species ISG15s.

References

1. ↑ Langley C, Goodwin O, Dzimianski JV, Daczkowski CM, Pegan SD. Structure of interferon-stimulated gene product 15 (ISG15) from the bat species Myotis davidii and the impact of interdomain ISG15 interactions on viral protein engagement. Acta Crystallogr D Struct Biol. 2019 Jan 1;75(Pt 1):21-31. doi:, 10.1107/S2059798318015322. Epub 2019 Jan 4. PMID:30644842 doi:http://dx.doi.org/10.1107/S2059798318015322
2. ↑ Narasimhan J, Wang M, Fu Z, Klein JM, Haas AL, Kim JJ. Crystal structure of the interferon-induced ubiquitin-like protein ISG15. J Biol Chem. 2005 Jul 22;280(29):27356-65. Epub 2005 May 24. PMID:15917233 doi:10.1074/jbc.M502814200
3. ↑ doi: https://dx.doi.org/10.2210/pdb5CHF/pdb