Spike (S) proteins of coronaviruses, including that which causes Severe Acute Respiratory Syndrome (SARS-CoV), associate with cellular receptors to mediate infection of their target cells. In Preliminary Data, we identify a metallopeptidase, angiotensin-converting enzyme 2 (ACE2), isolated from SARS-CoV-permissive Vero E6 cells, that efficiently binds the S1 domain of the SARS-CoV S protein. A soluble form of ACE2, but not that of a related enzyme, ACE1, blocked association of the S1 domain with Vero E6 cells. 293T cells transfected with ACE2, but not those transfected with HIV-1 receptors, formed syncytia with S-protein-expressing cells. Finally, SARS-CoV replicated efficiently on ACE2- but not mock-transfected 293T cells, and SARS-CoV replication could be blocked by anti-ACE2 but not anti-ACE1 antibodies. ACE2 is therefore a functional receptor for SARS-CoV. We propose to extend this work by (1) describing the role of ACE2 enzymatic activity in S-protein-mediated fusion; (2) assessing the ability of the S protein to modulate ACE2 activity; (3) describing the ability of various proteins, peptides, and small molecules that bind ACE2 to block S1 association, syncytia formation, and viral replication; (4) cloning ACE2 genes of a variety of animals, and describing their ability to associate with the S1 domain of the S protein and to support infection; (5) using chimeras of human and animal ACE2, and other ACE2 variants, to identify the S-protein-binding domain of ACE2; (6) identifying the minimum ACE2-binding domain of the S1 domain of the SARS-CoV S protein, and identifying residues within that domain essential for ACE2 association; and (7) optimizing an infection system using SIV pseudotyped with the S protein. These studies will provide a comprehensive description of the S-protein/ACE2 association and of the role of ACE2 in S-protein-mediated fusion.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
1R01AI061601-01
Application #
6816688
Study Section
Virology Study Section (VR)
Program Officer
Cassels, Frederick J
Project Start
2004-06-01
Project End
2004-12-31
Budget Start
2004-06-01
Budget End
2004-12-31
Support Year
1
Fiscal Year
2004
Total Cost
$136,008
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
030811269
City
Boston
State
MA
Country
United States
Zip Code
02115
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Li, Fang; Li, Wenhui; Farzan, Michael et al. (2005) Structure of SARS coronavirus spike receptor-binding domain complexed with receptor. Science 309:1864-8