2021 Fiscal Year Research-status Report
Real-time observation of native conformations and molecular behaviors of viral fusion proteins using high-speed atomic force microscopy (HS-AFM)
| Project/Area Number |
20K16262
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| Research Institution | Kanazawa University |
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Principal Investigator |
LIM KEE・SIANG 金沢大学, ナノ生命科学研究所, 特任助教 (60842987)
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| Project Period (FY) |
2020-04-01 – 2024-03-31
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| Keywords | SARS-CoV-2 spike protein / HS-AFM / Viral Fusion Protein / COVID-19 |
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| Outline of Annual Research Achievements |
The project is running smoothly. Due to COVID-19, spike (S) protein has been included in this project. HS-AFM imaging revealed the native structure of S protein. S protein underwent structural change at low pH, which the S1 subunit was dissociated and S2 subunit was elongated. Real-time observation of dynamic S protein-ACE2 interaction recorded the distinct S protein-ACE2 complex. Lastly, S protein or S2 subunit were used to interact with ACE2-expressing exosomes. Results showed two distinct interaction patterns: S protein bound to exosome via RBD-ACE2 interaction; S2 subunit used its fusion peptide to attach on exosome lipid layer and then drilled into the exosome. Altogether, molecular behavior of S protein at the nanoscopic level has been comprehensively studied using HS-AFM.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
Up to date, two important viral fusion proteins, Influenza A hemagglutinin (HA) and SARS-CoV-2 Spike (S) protein, have been comprehensively studied using HS-AFM. The results have been published in Nano Letter (Influenza A HA) and in Journal of Extracellular Vesicles (SARS-CoV-2 S protein). The achievements are motivating, and the project will be continued to finish the remaining viral fusion proteins.
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| Strategy for Future Research Activity |
The coming research direction focuses on fusion proteins of Dengue and Zika viruses. Experiments will be conducted to study their conformational dynamic.
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| Causes of Carryover |
Research grant will be used to purchase recombinant Dengue and Zika viral proteins, antibodies, and VLPs. In addition, consumables for HS-AFM imaging including mica, glass stage, and cantilevers will also be bought.
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