Mutagenesis analysis of the NS2B determinants of the Alkhurma virus NS2B-NS3 protease activation.
Alkhurma virus (ALKV) is a tick-borne class 4 flavivirus responsible for several human cases of haemorrhagic fever in Saudi Arabia, with no specific treatment currently available. The viral RNA encodes a serine protease (NS2B-NS3), essential for virus replication in infected cells, that constitutes an attractive target for antiviral compounds. In an attempt to identify residues and motifs on NS2B that are necessary for protease activity of the ALKV NS2B-NS3 complex, a series of modified NS2B-NS3 proteins was constructed, with point mutations on particular residues or with the NS2B domain derived from two different viruses. Four mutants and the two chimeric proteins exhibited reduction of protease activity against BAPNA (a p-nitroanilide substrate). The results demonstrate that tight complementarity of the protein sequences is necessary for NS2B-dependent activation of NS3. The results also determine residues in the ALKV NS2B cofactor essential for protease activation, giving new insights into protease function in flaviviruses.
Pastorino
Boris A M
BAM
Unité de Virologie Tropicale, Institut de Médecine Tropicale du Service de Santé des Armées (IMTSSA), BP 46, 13998 Marseille Armées, France.
Peyrefitte
Christophe N
CN
Unité de Virologie Tropicale, Institut de Médecine Tropicale du Service de Santé des Armées (IMTSSA), BP 46, 13998 Marseille Armées, France.
Grandadam
Marc
M
Unité de Virologie Tropicale, Institut de Médecine Tropicale du Service de Santé des Armées (IMTSSA), BP 46, 13998 Marseille Armées, France.
Thill
Maxime C E
MCE
Unité de Virologie Tropicale, Institut de Médecine Tropicale du Service de Santé des Armées (IMTSSA), BP 46, 13998 Marseille Armées, France.
Tolou
Hugues J
HJ
Unité de Virologie Tropicale, Institut de Médecine Tropicale du Service de Santé des Armées (IMTSSA), BP 46, 13998 Marseille Armées, France.
Bessaud
Maël
M
Unité de Virologie Tropicale, Institut de Médecine Tropicale du Service de Santé des Armées (IMTSSA), BP 46, 13998 Marseille Armées, France.
D000595:Amino Acid Sequence; D001586:Benzoylarginine Nitroanilide / Q000378:metabolism; D010450:Endopeptidases / Q000235:genetics*; D018067:Flaviviridae / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D008969:Molecular Sequence Data; D017354:Point Mutation; D016415:Sequence Alignment; D012697:Serine Endopeptidases / Q000378:metabolism*; D017361:Viral Nonstructural Proteins / Q000235:genetics*; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication
C080102:NS2B protein, flavivirus; D017361:Viral Nonstructural Proteins; D014764:Viral Proteins; D001586:Benzoylarginine Nitroanilide; D010450:Endopeptidases; D012697:Serine Endopeptidases