Investigation of ACE2 N-terminal fragments binding to SARS-CoV-2 Spike RBD

  1. G. Zhang
  2. S. Pomplun
  3. A. R. Loftis
  4. X. Tan
  5. A. Loas
  6. B. L. Pentelute

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Abstract

Coronavirus disease 19 (COVID-19) is an emerging global health crisis. With over 7 million confirmed cases to date, this pandemic continues to expand, spurring research to discover vaccines and therapies. SARS-CoV-2 is the novel coronavirus responsible for this disease. It initiates entry into human cells by binding to angiotensin-converting enzyme 2 (ACE2) via the receptor binding domain (RBD) of its spike protein (S). Disrupting the SARS-CoV-2-RBD binding to ACE2 with designer drugs has the potential to inhibit the virus from entering human cells, presenting a new modality for therapeutic intervention. Peptide-based binders are an attractive solution to inhibit the RBD-ACE2 interaction by adequately covering the extended protein contact interface. Using molecular dynamics simulations based on the recently solved cryo-EM structure of ACE2 in complex with SARS-CoV-2-RBD, we observed that the ACE2 peptidase domain (PD) α1 helix is important for binding SARS-CoV-2-RBD. Using automated fast-flow peptide synthesis, we chemically synthesized a 23-mer peptide fragment of the ACE2 PD α1 helix (SBP1) composed entirely of proteinogenic amino acids. Chemical synthesis of SBP1 was complete in 1.5 hours, and after work up and isolation >20 milligrams of pure material was obtained. Bio-layer interferometry (BLI) revealed that SBP1 associates with micromolar affinity to insect-derived SARS-CoV-2-RBD protein obtained from Sino Biological. Association of SBP1 was not observed to an appreciable extent to HEK cell-expressed SARS-CoV-2-RBD proteins and insect-derived variants acquired from other vendors. Moreover, competitive BLI assays showed SBP1 does not outcompete ACE2 binding to Sino Biological insect-derived SARS-CoV-2-RBD. Further investigations are ongoing to gain insight into the molecular and structural determinants of the variable binding behavior to different SARS-CoV-2-RBD protein variants.

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  1. Version published to 10.1101/2020.03.19.999318v2 on bioRxiv
  2. ScreenIT

    SciScore for 10.1101/2020.03.19.999318: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.Randomizationnot detected.Blindingnot detected.Power Analysisnot detected.Sex as a biological variablenot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    The association and dissociation curves were fitted with ForteBio Biosystems using four experimental conditions ( n = 4 , global fitting algorithm , binding model 1:1 ) to obtain the dissociation constant KD .
    ForteBio Biosystems
    suggested: None

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

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  3. Version published to 10.1101/2020.03.19.999318v1 on bioRxiv