In the ongoing coronavirus disease pandemic 2019 (COVID-19), many studies have focused on the role of neutralizing antibodies in fighting infection with severe acute coronavirus-2 respiratory syndrome (SARS-CoV-2). Most of these studies are based on genome sequencing, showing matches between the hypervariable region (HVR) and the antigen receptor binding domain (RBD). The results, however, are difficult to interpret due to many factors, such as the rapid decline in antibody titers, as well as the sharing of antigenic sequences with many other viruses such as earlier SARS (89% of common sequences) and influenza viruses.
New overprint on bioRxiv * the server describes the interactions between the SARS-CoV-2 virus spike binding protein and antibodies. Previous studies have dealt with similarities in the 3D structure of spike proteins in the two SARS viruses, as seen on cryo-EM. When comparing the amino acid sequences of the spike proteins of the two viruses, they showed 77% similarity. Because this shows no differences in binding, nor the influence of temperature and other factors, nor the nature of binding, these studies were not very successful.
The present study compared the binding strength of the SARS-CoV-2 and SARS-CoV protein-antibody complexes. Because viral interactions with its angiotensin receptor converting enzyme 2 (ACE2) or its IgM / IgG antibodies via a hydrogen bond between the carboxyl and amino groups are thought to have been used, the researchers attempted to use infrared spectroscopy (FTIR) to understand the nature of the bond as well. number of bonds. This would help in assessing the intensity of the viral attack.
Unusual temperature dependence
Surprisingly, they found almost the same number of bonds for any virus at room temperature or below 27 ° C. In other words, nonspecific antibody reactivity was observed, with an accuracy of less than 95%. When binding occurred at human body temperature, i.e., 37 ° C or above 31 ° C, antibody specificity improved according to the number of available hydrogen bonds, at 19 and 12, respectively.
This was due to the further development of the quaternary structure of the protein at higher temperatures. Thermal agitation leads to heat fracture of van der Waals bonds between protein strands, which reduces the number of such bonds. This led to the slow exposure of several hydrogen bonding sites.
FTIR showed similar absorbance at 1550 cm-1 for both mixtures, SARS-CoV-2 spike protein / SARS-CoV-2 antibody and SARS-CoV-2 112 spike protein / SARS-CoV-1 antibody, at 0.5% , or 0.4%. In other words, the bonds between the antibody and the spike protein were not strengthened, but a larger number of bonds formed at higher temperatures. This explains the greater specificity, as a higher binding ratio leads to effective blockade of binding to nonspecific antibodies.
These results are consistent with a recently published study confirming the number of hydrogen bonds in both virus-antibody complexes by amino acid sequencing and comparing the heavy chains of antibody molecules targeting both viruses.
Comparison of the amino acid sequence of antibody 2 and antibody 1 with respect to the difference in binding to the S protein. IgMs are shown by a blue band in the middle, and S proteins are represented by screw bands at the top and bottom, where some representative epitope residues in the S protein are indicated. The antibody sequence was obtained from the literature [32,33]. Both antibodies form only 2+ hydrogen bonds at room temperature due to folding of the protein structure, while such a number would increase to 19 and 12, respectively, at human body temperature. As can be calculated by the Arhenius equation, about 5% of the S protein would bind to antibody 1 at room temperature, resulting in the inevitable inaccuracy of the assay that could only be eliminated at body temperature.
What are the implications?
Despite the close phylogenetic bond and very similar sequences found in SARS-CoV and SARS-CoV-2, infrared spectroscopy showed the presence of a different number of hydrogen bonds between the corresponding ear molecules and antibodies. They found an increase in binding strength as a function of temperature, probably due to the unfolding of the protein quaternary structure. This has been found to be due to an increase in the binding site and thus the number of hydrogen bonds between the carboxyl and amino ends of different proteins.
Their findings were supported by absorbance findings at 37 ° C, when the number of hydrogen bonds of two types of antibodies to SARS-CoV-2 and SARS-CoV 19 versus 11 was found to be 11. At 27 ° C, the bond ratio calculated by thermodynamic exponents is about 20: 1, taking into account the non-specificity of binding at a lower temperature. Such information will help assess the accuracy of recent tests of rapid Covid-19 IgM / IgG antibodies, which are intended to help monitor the response to the vaccine more quickly and widely, as well as virus research in general.
Our results undoubtedly call for the need to simulate human body temperature in future antibody diagnoses, especially during the search for possible vaccines, because the uniqueness of binding or specificity of SARS-CoV-2 IgM / IgG could be fully available at warmer temperatures instead of conventional laboratory / room temperature of 20 ° C – 25 ° C, where most vaccine research is conducted. “
* Important notice
bioRxiv publishes preliminary scientific reports that have not been reviewed and should therefore not be considered final, guide clinical practice / health-related behavior, or treat it as established information.