A Natural Product Inhibitor of the West Nile Virus NS2B-NS3 Protease
Mentor:Nicholas Salzameda, Assistant Professor of Organic and Bioorganic Chemistry, California State University Fullerton
The West Nile Virus (WNV) is a mosquito-borne human sickness of the flavivirus genus and has been a global epidemic since the late 1990’s. Those infected with the WNV experience symptoms ranging from flu-like to long-term cognitive and neurological impairment, and even death. Currently, the only preventive measures for WNV are public awareness and mosquito control; there are no vaccines or other therapeutic treatments for WNV infections. A promising therapeutic target for the WNV is the WNV NS2B-NS3 protease, an enzyme that is vital for in vivo viral replication of the WNV. When this enzyme is inhibited, the WNV cannot replicate in cells, thereby halting the infection. Currently, there are no reported potent small molecule inhibitors for the WNV NS2B-NS3 protease. Therefore, our overall goal is to identify and optimize small molecule inhibitors for the WNV NS2B-NS3 protease. To accomplish this goal, we have created a novel FRET-based high throughput assay to screen small molecule libraries for WNV NS2B-NS3 protease inhibition. Our assay has excellent kinetic parameters, a Km of 3.35 ± 0.31 μM, a kcat of 0.0717 ± 0.0016 sec-1, and a kcat/ Km of 21,400 ± 2,000 M-1sec-1, and is a superb assay for the identification of true WNV NS2B-NS3 protease inhibitors. We utilized our FRET assay to screen the NIH clinical compound library for potential NS2B-NS3 protease inhibitors. This library is comprised of “old drugs” that have known therapeutic effects. We have subsequently identified a natural product from this library that inhibits the WNV NS2B-NS3 protease. Based on molecular modeling, our inhibitor is believed to bind in the active site of the NS2B-NS3 protease. Kinetic analysis shows an uncompetitive mode of inhibition with a Ki of 820 ± 90 nM.