(27aj) Artificial Lipid Biomembranes for Full-Length Sars-Cov-2 Receptor

The angiotensin-converting enzyme 2 (ACE2), as a functional receptor for SARS-CoV-2, is essential for preliminary assessment of potential hosts and treatment strategies. However, many studies are based on its truncated version but not full-length structure. Compared to the truncated version, a single-transmembrane helix presents in the full-length structure of ACE2, influencing its interaction and binding with SARS-CoV-2. Therefore, synthesis of the full-length ACE2 is an urgent requirement. Here, cell-free membrane protein synthesis systems (CFMPSs) are constructed for full-length membrane proteins. The transmembrane protein MscL is screened as a model among 10 membrane proteins based on their expression and solubility. Next, CFMPSs are constructed and optimized based on natural vesicles, vesicles with outer 4 membrane proteins removed or 2 chaperonins added, and 37 types of nanodiscs. They all increase membrane protein solubility to more than 50%. Finally, the full-length ACE2 of 21 species are successfully expressed with yields between 0.4-0.9 mg/mL using the constructed CFMPS. Their functions show definite differences from the truncated versions, suggesting that the transmembrane region of ACE2 affects its structure and function. CFMPSs can be further extended to a broader range of membrane proteins, paving the way for their use in basic and applied research.