Dimeric ACE2-FC is equivalent to monomeric ACE2 in the surrogate virus neutralization test

Abstract

Angiotensin converting enzyme 2 (ACE2) is the main cellular receptor for the dangerous sarbecoviruses SARS-CoV and SARS-CoV-2. Its recombinant extracellular domain is used to monitor the level of the protective humoral immune response to a viral infection or vaccine using a surrogate virus neutralization test (sVNT). Soluble ACE2 is also being considered as an antiviral therapy option potentially insensitive to changes in the SARS-CoV-2 spike protein. For widespread sVHT testing, it is necessary to use ACE2 preparations or ACE2 conjugates with constant properties. Previously, we obtained a cell line that produces soluble monomeric ACE2 and showed that this variant of ACE2 can be used in sBHT, preferably in the form of a conjugate with horseradish peroxidase. To obtain a stable and universally applicable form of soluble ACE2, a cell line was obtained that produced the ACE2-Fc fusion protein with high productivity, more than 150 mg/l of the target protein during cultivation in a stirred flask. Affinity-purified ACE2-Fc is a mixture of dimeric and tetrameric forms, but allows one to obtain linearizable antibody inhibition curves for complexation with the receptor-binding domain of the SARS-CoV-2 spike protein. The ACE2-Fc-HRP based sVHT testing system can be used to practically measure the levels of virus-neutralizing antibodies against various circulating variants of the SARS-CoV-2 virus.