NS3 protease of Langat tick-borne flavivirus cleaves serine protease substrates.
Langat (LGT) virus, initially isolated in 1956 from ticks in Malaysia, is a naturally occurring nonpathogenic virus with a very close antigenicity to the highly pathogenic tick-borne encephalitis (TBE) Western subtype virus and TBE Far Eastern subtype virus. NS3, the second largest viral protein of LGT virus, is highly conserved among flaviviruses and contains a characteristic protease moiety (NS3 pro). NS3 pro represents an attractive target for anti-protease molecules against TBE virus. We report herein a purification method specially designed for NS3 pro of LGT using a strategy for proper refolding coupled with the enzymatic characterisation of the protein. Different p-nitroanilide substrates, defined on canonic sequences for their susceptibility to Ser-protease, were applied to the proteolytic assays of the protein. The highest values were obtained from substrates containing an Arg or Lys (amino acid) residue at the P1 position. This purification method will facilitate the future development of reliable testing procedures for anti-proteases directed to NS3 proteins.
Scaramozzino
Natale
N
Unité de Virologie, Centre de Recherches du Service de Santé des Armées Emile, Pardé, F-38000 Grenoble, France.
Crance
Jean-Marc
JM
Drouet
Christian
C
Roebuck
Jean-Philippe
JP
Drouet
Emmanuel
E
Jouan
Alain
A
Garin
Daniel
D
D000595:Amino Acid Sequence; D000818:Animals; D002863:Chromogenic Compounds / Q000378:metabolism; D004669:Encephalitis Viruses, Tick-Borne / Q000201:enzymology*; D004675:Encephalitis, Tick-Borne / Q000201:enzymology / Q000821:virology; D008296:Malaysia; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000627:therapeutic use; D017510:Protein Folding; D020365:RNA Helicases; D016415:Sequence Alignment; D012697:Serine Endopeptidases / Q000378:metabolism*; D013987:Ticks; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D002863:Chromogenic Compounds; D011480:Protease Inhibitors; D020365:RNA Helicases; D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; C060028:NS3 protein, flavivirus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases