ABSTRACT
The COVID-19 pandemic has significantly impacted human health for three years. To mitigate the spread of SARS-CoV-2, the development of neutralizing antibodies has been accelerated, including the exploration of alternative antibody formats such as single-domain antibodies. In this study, we identified variable new antigen receptors (VNARs) specific for the receptor binding domain (RBD) of SARS-CoV-2 by immunizing a banded houndshark (Triakis scyllium) with recombinant wild-type RBD. Notably, the CoV2NAR-1 clone showed high binding affinities in the nanomolar range to various RBDs and demonstrated neutralizing activity against SARS-CoV-2 pseudoviruses. These results highlight the potential of the banded houndshark as an animal model for the development of VNAR-based therapeutics or diagnostics against future pandemics.
Subject(s)
COVID-19 , Single-Domain Antibodies , Humans , Animals , SARS-CoV-2/metabolism , Antibodies, Viral , Pandemics , Antibodies, NeutralizingABSTRACT
A new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant known as Omicron has caused a rapid increase in recent global patients with coronavirus infectious disease 2019 (COVID-19). To overcome the COVID-19 Omicron variant, production of a recombinant spike receptor binding domain (RBD) is vital for developing a subunit vaccine or a neutralizing antibody. Although bacterial expression has many advantages in the production of recombinant proteins, the spike RBD expressed in a bacterial system experiences a folding problem related to disulfide bond formation. In this study, the soluble Omicron RBD was obtained by a disulfide isomerase-assisted periplasmic expression system in Escherichia coli. The Omicron RBD purified from E. coli was very well recognized by anti-SARS-CoV-2 antibodies, sotrovimab (S309), and CR3022, which were previously reported to bind to various SARS-CoV-2 variants. In addition, the kinetic parameters of the purified Omicron RBD upon binding to the human angiotensin-converting enzyme 2 (ACE2) were similar to those of the Omicron RBD produced in the mammalian expression system. These results suggest that an E. coli expression system would be suitable to produce functional and correctly folded spike RBDs of the next emerging SARS-CoV-2 variants quickly and inexpensively.
ABSTRACT
The frequent occurrence of viral variants is a critical problem in developing antiviral prophylaxis and therapy; along with stronger recognition of host cell receptors, the variants evade the immune system-based vaccines and neutralizing agents more easily. In this work, we focus on enhanced receptor binding of viral variants and demonstrate generation of receptor-mimicking synthetic reagents, capable of strongly interacting with viruses and their variants. The hotspot interaction of viruses with receptor-derived short peptides is maximized by aptamer-like scaffolds, the compact and stable architectures of which can be in vitro selected from a myriad of the hotspot peptide-coupled random nucleic acids. We successfully created the human angiotensin-converting enzyme 2 (hACE2) receptor-mimicking hybrid ligand that recruits the hACE2-derived receptor binding domain-interacting peptide to directly interact with a binding hotspot of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Experiencing affinity boosting by ~500% to Omicron, the de novo selected hACE2 mimic exhibited a great binding tolerance to all SARS-CoV-2 variants of concern.