Screening and characterization of inhibitory vNAR targeting nanodisc-assembled influenza M2 proteins
Chuandi Yu1,2, Wen Ding3, Lei Zhu1
1High Magnetic Field Laboratory, Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei, Anhui 230031, China.
Abstract:
Influenza A virus poses a constant challenge to human health. The highly conserved influenza matrix-2 (M2) protein is an attractive target for the development of a universal antibody-based drug. However, screening using antigens with subphysiological conformation in a nonmembrane environment significantly reduces the generation of efficient antibodies. Here, M2(1-46) was incorporated into nanodiscs (M2-nanodiscs) with M2 in a membrane-embedded tetrameric conformation, closely resembling its natural physiological state in the influenza viral envelope. M2-nanodisc generation, an antigen, was followed by Chiloscyllium plagiosum immunization. The functional vNARs were selected by phage display panning strategy from the shark immune library. One of the isolated vNARs, AM2H10, could specifically bind to tetrameric M2 instead of monomeric M2e (the ectodomain of M2 protein). Furthermore, AM2H10 blocked ion influx through amantadine-sensitive and resistant M2 channels. Our findings indicated the possibility of developing functional shark nanobodies against various influenza viruses by targeting the M2 protein.
Insights
Researchers developed functional shark nanobodies targeting the influenza A virus M2 protein. These nanobodies, derived from M2-nanodiscs, effectively block viral ion channels, offering a potential universal antibody-based drug against influenza.
Area of Science:
- Virology
- Immunology
- Biochemistry
Background:
- Influenza A virus presents a continuous global health threat.
- The influenza matrix-2 (M2) protein is a conserved target for universal antiviral therapies.
- Generating effective antibodies against M2 is challenging due to conformational issues in non-membrane environments.
Purpose of the Study:
- To develop functional antibody-based therapeutics against influenza A virus.
- To generate antibodies targeting the M2 protein in its native tetrameric conformation.
- To explore the potential of shark nanobodies (vNARs) for influenza therapy.
Main Methods:
- Incorporation of M2(1-46) into nanodiscs (M2-nanodiscs) to mimic physiological conformation.
- Immunization of sharks (Chiloscyllium plagiosum) with M2-nanodiscs.
- Phage display panning of a shark immune library to select functional vNARs.
- Characterization of vNAR binding specificity and functional activity.
Main Results:
- Successfully generated M2-nanodiscs presenting M2 in a membrane-embedded tetrameric conformation.
- Isolated a specific vNAR, AM2H10, that binds selectively to tetrameric M2, not monomeric M2e.
- AM2H10 demonstrated the ability to block ion flux through M2 channels, including amantadine-resistant variants.
Conclusions:
- M2-nanodiscs are effective antigens for generating conformationally relevant antibodies.
- Shark nanobodies, like AM2H10, can be developed into functional inhibitors of the influenza M2 channel.
- This approach shows promise for creating universal antibody-based drugs against diverse influenza A viruses.
