Project Grant R21AI183195

STRUCTURE-FUNCTION ANALYSIS OF HUMAN AND BAT CORONAVIRUS SPIKE PROTEINS - PROJECT SUMMARY/ABSTRACT THE COVID-19 PANDEMIC HAS AFFECTED OVER 690 MILLION PEOPLE WORLDWIDE AND CAUSED 6.9 MILLION DEATHS. ALTHOUGH THE RECENT VARIANTS ARE LESS INFECTIOUS AND CAUSE LOWER MORTALITY THAN EARLIER ONES, DIAGNOSED ACTIVE CASES ARE AS HIGH AS 20 MILLION WORLDWIDE. SARS-COV-2 SPIKE PROTEIN IS CONTINUOUSLY ACCUMULATING MUTATIONS. OUR EARLIER PUBLISHED STUDIES ON ITS RECEPTOR-BINDING DOMAIN (RBD) SHOWED THAT INCREASED RECEPTOR BINDING AND ESCAPE FROM NEUTRALIZING ANTIBODIES DIRECT THE NATURAL SELECTION OF MUTATIONS. HOWEVER, MUTATIONS HAVE ALSO BEEN FOUND IN OTHER REGIONS OF THE SPIKE PROTEIN, AND THE ROLE OF THESE NON-RBD MUTATIONS IS NOT WELL UNDERSTOOD. COMPUTATIONAL MODELS PROPOSE A LONG-RANGE ALLOSTERIC COMMUNICATION BETWEEN DIFFERENT PARTS OF THE SPIKE PROTEIN. DURING THIS 2-YEAR PROPOSAL PERIOD, WE WILL DESIGN EXPERIMENTS TO TEST THE ROLE OF LONG-RANGE COMMUNICATION BY PROBING THE EFFECT OF DISTANT MUTATIONS ON RBD BINDING TO ACE2 AND TO NEUTRALIZING ANTIBODIES (AIM 1). SPIKE PROTEIN IS A TRIMER AND EXISTS IN MULTIPLE CONFORMATIONS WITH INDIVIDUAL RBDS EITHER ACCESSIBLE OR INACCESSIBLE TO ACE2 AND ANTIBODIES. WE WILL PROBE THE RELATIVE POPULATIONS OF THE FOUR CONFORMATIONAL STATES OF THE SPIKE PROTEIN AND THE EFFECT OF MUTATIONS. IN ADDITION TO SARS-COV-2 EMERGED IN 2019, TWO OTHER CORONAVIRUSES (COVS) HAVE CAUSED SEVERE DISEASE: SARS-COV EMERGED IN 2003 AND MERS- COV IN 2012. THOUGH BASED ON ONLY THREE DATA POINTS, THE DATA SUGGESTS ANOTHER COVID OUTBREAK MIGHT BE IMMINENT WITHIN THE NEXT TEN YEARS. HUMAN CORONAVIRUSES (COVS) ARE WIDELY CONSIDERED TO HAVE ORIGINATED FROM BATS, AND HENCE, MULTIPLE EFFORTS WORLDWIDE HAVE BEEN FOCUSED ON IDENTIFYING BAT COVS CAPABLE OF INFECTING HUMANS. SO FAR, ABOUT 700 BAT COVS WITH UNIQUE SPIKE PROTEINS HAVE BEEN IDENTIFIED, AND IT IS CRUCIAL TO DETERMINE WHICH RECEPTOR PATHWAYS THEY MIGHT USE TO INFECT HUMANS IN ORDER TO DEVELOP EFFECTIVE THERAPIES. TO BEGIN WITH, WE WILL EXAMINE WHICH OF THESE BAT COV SPIKE PROTEINS MIGHT USE THE ACE2 (SARS-COV/SARS- COV-2) AND DPP4 (MERS-COV) PATHWAYS BY COMBINING BIOINFORMATIC SEQUENCE ANALYSIS AND DETERMINING THE RELATIVE BINDING AFFINITIES (AIM 2). THESE ASSAYS WILL BE FURTHER APPLIED TO DETERMINE WHICH OTHER RECEPTOR PATHWAYS BAT COVS MIGHT USE TO INFECT THE HUMAN RESPIRATORY SYSTEM. THE MOLECULAR KNOWLEDGE GAINED DURING THIS PROPOSAL WILL HELP US IN PREDICTING THE EVOLUTION OF THE CURRENT SARS-COV-2 VIRUS AND ANY FUTURE COVID OUTBREAKS, WHICH WILL FINALLY LEAD TO DEVELOPING EFFECTIVE THERAPEUTICS THAT CAN NEUTRALIZE MULTIPLE COVS.