Diversity of ACE 2 and its interaction with SARS-CoV-2 receptor binding domain 1 2

12 COVID-19, the clinical syndrome caused by the SARS-CoV-2 virus, has rapidly spread globally causing 13 hundreds of millions of infections and over two million deaths. The potential animal reservoirs for 14 SARS-CoV-2 are currently unknown, however sequence analysis has provided plausible potential 15 candidate species. SARS-CoV-2 binds to the angiotensin I converting enzyme 2 (ACE2) to enable its 16 entry into host cells and establish infection. We analyzed the binding surface of ACE2 from several 17 important animal species to begin to understand the parameters for the ACE2 recognition by the SARS18 CoV-2 spike protein receptor binding domain (RBD). We employed Shannon entropy analysis to 19 determine the variability of ACE2 across its sequence and particularly in its RBD interacting region, and 20 assessed differences between various species’ ACE2 and human ACE2. Recombinant ACE2 from 21 human, hamster, horseshoe bat, cat, ferret, and cow were evaluated for RBD binding. A gradient of 22 binding affinities were seen where human and hamster ACE2 were similarly in the low nanomolar 23 range, followed by cat and cow. Surprisingly, horseshoe bat (Rhinolophus sinicus) and ferret (Mustela 24 putorius) ACE2s had poor binding activity compared to the other species’ ACE2. The residue 25 differences and binding properties between the species’ variants provide a framework for 26 understanding ACE2-RBD binding and virus tropism. 27