Structure-guided design of a CD81-binding mini-protein that blocks hepatitis C virus entry

Authors

Cho, H.-S., An, S., Jo, E., Choi, H. S., Kim, S. S., Kim, D. W., Yang, W., Jang, S. K., Park, K.-H. P., Ha, S.-J., Gho, Y. S., Yang, S. W., Kim, S.

Abstract

Despite the success of direct-acting antivirals, preventing hepatitis C virus (HCV) reinfection remains a critical global challenge. To address this, we leveraged deep learning-based de novo protein design to engineer mini-proteins targeting the large extracellular loop (LEL) of the HCV co-receptor CD81. These mini-proteins are predicted to precisely dock into CD81-LEL, occluding the critical binding interface required for the HCV E2 glycoprotein. Through biophysical screening, we identified mini-19, a lead candidate with sub-nanomolar affinity (KD = 0.5 nM) and high thermostability up to 95{degrees}C. Flow cytometry and super-resolution imaging confirmed that mini-19 specifically recognizes the native topology of CD81 on cells and extracellular vesicles. Functionally, mini-19 neutralized HCVcc infection (IC50 = 1.2 nM). Molecular dynamics simulations demonstrated that mini-19 acts as a structural clamp, restricting the fusogenic conformational plasticity of the receptor. By targeting a conserved host factor rather than the rapidly mutating viral envelope, mini-19 provides a high genetic barrier to resistance. Beyond offering a prophylactic strategy against HCV, our findings establish a stable biologic platform for targeting tetraspanin-enriched microdomains and modulating host-pathogen interactions.

Preprint server: bioRxiv
The authors list and abstract were imported from bioRxiv on 14 Jul 2026.

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