Journal: bioRxiv

Article Title: Binding strength and hydrogen bond numbers between Covid-19 RBD and HVR of antibody

doi: 10.1101/2020.12.21.423787

Figure Lengend Snippet: (a) Schematic illustration of the hydrogen bonding between carboxyl and amino groups of the protein strands, representing a typical intermolecular binding for the secondary structure of proteins. (b) The FTIR apparatus, the ATR model has been adopted to improve the signal noise ratio, which offers 0.1% error margin in the absorption measurement, and 0.5cm -1 along the wavenumber scan. To facilitate the possible temperature variation, an electric heating element, and a thermometer were attached to the metallic plate of the ATR assembly, which also acts as an ideal heat sink. Samples of antibodies and spike proteins were mixed together, and undergone FTIR spectroscopic examination conditioned by variable temperatures. (c) The FTIR scan of the sample mixture of SARS-CoV-2 spike protein/ SARS-CoV-2 antibody (S + Antibody 2) in PBS solution. Comparing the FTIR signal of the PBS solvent, it indicates that 1550 cm -1 was the only absorption band exclusively associated with the proteins, but not overshadowed by the PBS solvent. (d) The same FTIR scan of the sample mixture of SARS-CoV-2 spike protein/ SARS-CoV-1 antibody (S + Antibody 1) in PBS solution.

Article Snippet: SARS-CoV-2 Spike RBD-His (V367F) Recombinant Protein (Purity: > 95 % as determined by SDS-Page, protein concentration: 0.1wt%), SARS-CoV-2 antibody (antibody 2, also named SARS-CoV-2 (2019-nCoV) Spike Neutralizing Antibody), and SARS-CoV-1 antibody (antibody 1, also named SARS-CoV/SARS-CoV-2 Spike S1 antibody) (Purity: > 95 % as determined by SDS-Page, protein concentration: 0.1wt%) were all purchased from Sino Biological Inc. PBS solution (phosphate buffered solution containing 0.1wt%NaCl +0.002wt%KCl +0.01wt%Na 2 HPO 4 +0.002wt%KH 2 PO 4 +99 + wt%H 2 O) was purchased from Solarbio Corp. A Nicolet iS50 FT-IR spectrometer was adopted to acquire the FTIR spectra, in wavenumber range of 500-4000 cm -1 by the attenuated total reflectance (ATR) mode.

Techniques: Binding Assay

Journal: bioRxiv

Article Title: Binding strength and hydrogen bond numbers between Covid-19 RBD and HVR of antibody

doi: 10.1101/2020.12.21.423787

Figure Lengend Snippet: (a) Detailed FTIR scan of the sample mixture of SARS-CoV-2 spike protein/ SARS-CoV-2 antibody (S + Antibody 2) in PBS solution, within the wavenumber range of 1400-1800cm -1 . (b) Detailed FTIR scan of the sample mixture of SARS-CoV-2 spike protein/ SARS-CoV-1 antibody (S + Antibody 1) in PBS solution, in the wavenumber range of 1400-1800cm -1 . (c) The infrared absorptions of 1550cm -1 under variable temperature of the sample combination of SARS-CoV-2 spike protein/ SARS-CoV-2 antibody, versus the sample combination of SARS-CoV-2 spike protein/ SARS-CoV-1 antibody, obtained from the difference between the peak near 1550cm -1 and the trough near 1480cm -1 , after subtracting that of the buffer solution. The associated error bars were estimated by combining the instrumental error, and the possible deviation involved in the computation. A strong temperature dependence is observed, where the bonding number of the antibody is enhanced sharply beyond 31°C, rather than at the usual room temperature. (d) The possible temperature influence on the structure of spike protein and antibodies, where only the top part of the IgM, similar to IgG, was depicted: a higher temperature increases the probability of protein quaternary structure unfolding, and exposes more binding sites, hence more bonds.

Article Snippet: SARS-CoV-2 Spike RBD-His (V367F) Recombinant Protein (Purity: > 95 % as determined by SDS-Page, protein concentration: 0.1wt%), SARS-CoV-2 antibody (antibody 2, also named SARS-CoV-2 (2019-nCoV) Spike Neutralizing Antibody), and SARS-CoV-1 antibody (antibody 1, also named SARS-CoV/SARS-CoV-2 Spike S1 antibody) (Purity: > 95 % as determined by SDS-Page, protein concentration: 0.1wt%) were all purchased from Sino Biological Inc. PBS solution (phosphate buffered solution containing 0.1wt%NaCl +0.002wt%KCl +0.01wt%Na 2 HPO 4 +0.002wt%KH 2 PO 4 +99 + wt%H 2 O) was purchased from Solarbio Corp. A Nicolet iS50 FT-IR spectrometer was adopted to acquire the FTIR spectra, in wavenumber range of 500-4000 cm -1 by the attenuated total reflectance (ATR) mode.

Techniques: Binding Assay

Journal: bioRxiv

Article Title: Binding strength and hydrogen bond numbers between Covid-19 RBD and HVR of antibody

doi: 10.1101/2020.12.21.423787

Figure Lengend Snippet: (a) The FTIR scan of SARS-CoV-2 spike protein (S protein) in PBS solution, for the wavenumber range of 1400-1800cm -1 . (b) The FTIR scan of SARS-CoV-2 antibody (antibody 2) in PBS solution, for the wavenumber range of 1400-1800cm -1 . (c) The FTIR scan of SARS-CoV-1 antibody (antibody 1) in PBS solution, for the wavenumber range of 1400-1800cm -1 . (d) The infrared absorption of 1550cm -1 under variable temperature, of SARS-CoV-2 spike protein, SARS-CoV-2 antibody, and SARS-CoV-1 antibody, respectively, given by the difference between the crest near 1550cm -1 , and the trough near 1480cm -1 . The associated error bars were estimated by combining the instrumental error, and the possible deviation involved in the computation. All 3 proteins followed a similar trend in temperature, which is attributed to the thermal agitation, when the likelihood increases for such binding sites to pair, either between the neighbouring proteins, or amongst various strands within a protein, or even along a protein strand. (e) Schematic illustration of the hydrogen bonding between carboxyl and amino groups within a protein strand, showing the intra-molecular binding of the quaternary structure of proteins, when the Van der Waals bonds between large pendant groups, such as phenyl, were disrupted, which causes unfolding and exposing more bonding sites.

Article Snippet: SARS-CoV-2 Spike RBD-His (V367F) Recombinant Protein (Purity: > 95 % as determined by SDS-Page, protein concentration: 0.1wt%), SARS-CoV-2 antibody (antibody 2, also named SARS-CoV-2 (2019-nCoV) Spike Neutralizing Antibody), and SARS-CoV-1 antibody (antibody 1, also named SARS-CoV/SARS-CoV-2 Spike S1 antibody) (Purity: > 95 % as determined by SDS-Page, protein concentration: 0.1wt%) were all purchased from Sino Biological Inc. PBS solution (phosphate buffered solution containing 0.1wt%NaCl +0.002wt%KCl +0.01wt%Na 2 HPO 4 +0.002wt%KH 2 PO 4 +99 + wt%H 2 O) was purchased from Solarbio Corp. A Nicolet iS50 FT-IR spectrometer was adopted to acquire the FTIR spectra, in wavenumber range of 500-4000 cm -1 by the attenuated total reflectance (ATR) mode.

Techniques: Binding Assay