title: Mutagenesis of the West Nile virus NS2B cofactor domain reveals two regions essential for protease activity.
abstract: The flavivirus NS2B/NS3 protease has received considerable attention as a target for the development of antiviral compounds. While substrate based inhibitors have been the primary focus to date, an approach focussing on NS2B cofactor displacement could prove to be an effective alternative. To understand better the role of the NS2B cofactor in protease activation, we conducted an alanine mutagenesis screen throughout the 42-residue central cofactor domain (NS2B(51-92)) of West Nile virus (WNV). Two sites critical for proteolytic activity were identified (NS2B(59-62) and NS2B(75-87)), where the majority of substitutions were found to significantly decrease proteolytic activity of a recombinant WNV NS2B/NS3 protease. These findings provide mechanistic insights into the structural and functional role that the cofactor may play in the substrate-bound and free protease complexes as well as providing novel sites for targeting new antiviral inhibitors.
authors.affiliation: School of Molecular and Microbial Sciences, University of Queensland, Brisbane, QLD 4072, Australia.
authors.forename: Keith J
authors.initials: KJ
authors.lastname: Chappell
authors.affiliation: Institute for Molecular Bioscience, University of Queensland, Brisbane, QLD 4072, Australia.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Stoermer
authors.affiliation: Institute for Molecular Bioscience, University of Queensland, Brisbane, QLD 4072, Australia.
authors.forename: David P
authors.initials: DP
authors.lastname: Fairlie
authors.affiliation: Institute for Molecular Bioscience, University of Queensland, Brisbane, QLD 4072, Australia.|School of Molecular and Microbial Sciences, University of Queensland, Brisbane, QLD 4072, Australia.
authors.forename: Paul R
authors.initials: PR
authors.lastname: Young
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain; D003067:Coenzymes; D010450:Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018177:Flavivirus Infections / Q000821:virology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014901:West Nile Fever; D014902:West Nile virus / Q000737:chemistry*
chemical_list: D003067:Coenzymes; C080102:NS2B protein, flavivirus; D017361:Viral Nonstructural Proteins; D010450:Endopeptidases