Selection, biophysical and structural analysis of synthetic nanobodies that effectively neutralize SARS-CoV-2

Custodio T, Das H, Sheward D, Hanke L, Pazicky S, Pieprzyk J, Sorgenfrei M, Schroer M, Gruzinov A, Jeffries C, Graewert M, Svergun D, Dobrev N, Remans K, Seeger M, McInerney G, Murrell B, Hällberg B, Löw C, (2020) DOI

SASDJH4 – SARS-CoV-2 spike protein ACE2 receptor binding domain (RBD) bound to the synthetic nanobody Sybody 23 (Sy23)

Synthetic nanobody Sybody 23
Spike glycoprotein (ACE2 receptor binding domain)
MWI(0) 63 kDa
MWexpected 45 kDa
VPorod 87 nm3
log I(s) 5.30×10-2 5.30×10-3 5.30×10-4 5.30×10-5
Synthetic nanobody Sybody 23 Spike glycoprotein (ACE2 receptor binding domain) small angle scattering data  s, nm-1
ln I(s)
Synthetic nanobody Sybody 23 Spike glycoprotein (ACE2 receptor binding domain) Guinier plot ln 5.30×10-2 Rg: 3.5 nm 0 (3.5 nm)-2 s2
(sRg)2I(s)/I(0)
Synthetic nanobody Sybody 23 Spike glycoprotein (ACE2 receptor binding domain) Kratky plot 1.104 0 3 sRg
p(r)
Synthetic nanobody Sybody 23 Spike glycoprotein (ACE2 receptor binding domain) pair distance distribution function Rg: 3.7 nm 0 Dmax: 15.1 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Synthetic nanobody Sybody 23 Spike glycoprotein (ACE2 receptor binding domain) CORAL model

Synchrotron SAXS data from solutions of SARS-CoV-2 spike protein ACE2 receptor binding domain (RBD) bound to the synthetic nanobody Sybody 23 (Sy23) in 25 mM Tris 100 mM NaCl, pH 7.5 were collected on the EMBL P12 beam line at the PETRA III storage ring (Hamburg, Germany) using a Pilatus 6M detector at a sample-detector distance of 3 m and at a wavelength of λ = 0.124 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 2.80 mg/ml was measured at 20°C. 10 successive 0.100 second frames were collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

Synthetic nanobody Sybody 23 (Sy23)
Mol. type   Protein
Organism   synthetic construct
Olig. state   Monomer
Mon. MW   15.7 kDa
Sequence   FASTA
 
Spike glycoprotein (ACE2 receptor binding domain) (RBD SARS-CoV-2)
Mol. type   Protein
Organism   Severe acute respiratory syndrome coronavirus 2
Olig. state   Monomer
Mon. MW   29.3 kDa
 
UniProt   P0DTC2
Sequence   FASTA