Author : Luíza Mamigonian Bessa
Publisher :
ISBN 13 :
Total Pages : 0 pages
Book Rating : 4.:/5 (124 download)
Book Synopsis Investigation of the Hepatitis C Virus RNA Polymerase NS5B in Solution by Nuclear Magnetic Resonance and Its Interaction with Intrinsically Disordered Domain 2 of the NS5A Protein by : Luíza Mamigonian Bessa
Download or read book Investigation of the Hepatitis C Virus RNA Polymerase NS5B in Solution by Nuclear Magnetic Resonance and Its Interaction with Intrinsically Disordered Domain 2 of the NS5A Protein written by Luíza Mamigonian Bessa and published by . This book was released on 2017 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: NS5B is the hepatitis C virus (HCV) RNA-dependent RNA polymerase. This protein has been extensively studied by X-ray crystallography and shows an organization in three subdomains called fingers, palm and thumb. Whereas static crystallographic data are abundant, structural studies of this protein in solution are limited. Nuclear magnetic resonance (NMR) spectroscopy was used to study the 65 kDa NS5B in solution as well as its interaction with binding partners. It was characterized using selective isotopic labeling of isoleucine side-chain methyl groups, which gives rise to a simplified NMR spectrum with an improved signal-to-noise ratio. This characterization confirmed the presence of particular dynamics in the subdomains, especially in the thumb, as well as long-range effects that are transmitted through to other subdomains. Furthermore, this system was used to investigate the binding of the domain 2 of NS5A (NS5A-D2), a disordered domain of another HCV protein that has been shown to directly interact with NS5B in vitro. With paramagnetic relaxation enhancement experiments we showed that NS5A-D2 binds to NS5B via, at least, two binding sites on the thumb subdomain. As one of these sites was the binding site of allosteric inhibitor filibuvir, we characterized the binding of this small molecule to NS5B by NMR and found long-range effects of its binding throughout the polymerase. Finally, we studied the binding of a small RNA template strand to NS5B and found that both NS5A-D2 and filibuvir reduce but do not abolish the interaction between the polymerase and RNA. In sum, NMR spectroscopy was used to study dynamic properties of NS5B and its interactions with binding partners.