8DT8

LM18/Nb136 bispecific tetra-nanobody immunoglobulin in complex with SARS-CoV-2-6P-Mut7 S protein (focused refinement)

Summary for 8DT8

• Entry DOI: 10.2210/pdb8dt8/pdb
• EMDB information: 27692
• Descriptor: Spike glycoprotein, LM18 nanobody, Nb136 nanobody, ... (4 entities in total)
• Functional Keywords: nanobody, bispecific nanobody, coronavirus, antibody engineering, viral protein
• Biological source: Severe acute respiratory syndrome coronavirus 2; More
• Total number of polymer chains: 5
• Total formula weight: 454536.04

Authors

Ozorowski, G.,Turner, H.L.,Ward, A.B. (deposition date: 2022-07-25, release date: 2023-06-14, Last modification date: 2024-11-20)

Primary citation

Misson Mindrebo, L.,Liu, H.,Ozorowski, G.,Tran, Q.,Woehl, J.,Khalek, I.,Smith, J.M.,Barman, S.,Zhao, F.,Keating, C.,Limbo, O.,Verma, M.,Liu, J.,Stanfield, R.L.,Zhu, X.,Turner, H.L.,Sok, D.,Huang, P.S.,Burton, D.R.,Ward, A.B.,Wilson, I.A.,Jardine, J.G.
Fully synthetic platform to rapidly generate tetravalent bispecific nanobody-based immunoglobulins.
Proc.Natl.Acad.Sci.USA, 120:e2216612120-e2216612120, 2023

PubMed Abstract: Nanobodies bind a target antigen with a kinetic profile similar to a conventional antibody, but exist as a single heavy chain domain that can be readily multimerized to engage antigen via multiple interactions. Presently, most nanobodies are produced by immunizing camelids; however, platforms for animal-free production are growing in popularity. Here, we describe the development of a fully synthetic nanobody library based on an engineered human V3-23 variable gene and a multispecific antibody-like format designed for biparatopic target engagement. To validate our library, we selected nanobodies against the SARS-CoV-2 receptor-binding domain and employed an on-yeast epitope binning strategy to rapidly map the specificities of the selected nanobodies. We then generated antibody-like molecules by replacing the V and V domains of a conventional antibody with two different nanobodies, designed as a molecular clamp to engage the receptor-binding domain biparatopically. The resulting bispecific tetra-nanobody immunoglobulins neutralized diverse SARS-CoV-2 variants with potencies similar to antibodies isolated from convalescent donors. Subsequent biochemical analyses confirmed the accuracy of the on-yeast epitope binning and structures of both individual nanobodies, and a tetra-nanobody immunoglobulin revealed that the intended mode of interaction had been achieved. This overall workflow is applicable to nearly any protein target and provides a blueprint for a modular workflow for the development of multispecific molecules.

PubMed: 37276407
DOI: 10.1073/pnas.2216612120

Experimental method

ELECTRON MICROSCOPY (3.34 Å)