Cleavage targets and the D-arginine-based inhibitors of the West Nile virus NS3 processing proteinase.
Mosquito-borne WNV (West Nile virus) is an emerging global threat. The NS3 proteinase, which is essential for the proteolytic processing of the viral polyprotein precursor, is a promising drug target. We have isolated and biochemically characterized the recombinant, highly active NS3 proteinase. We have determined that the NS3 proteinase functions in a manner that is distantly similar to furin in cleaving the peptide and protein substrates. We determined that aprotinin and D-arginine-based 9-12-mer peptides are potent inhibitors of WNV NS3 with K(i) values of 26 nM and 1 nM respectively. Consistent with the essential role of NS3 activity in the life cycle of WNV and with the sensitivity of NS3 activity to the D-arginine-based peptides, we showed that nona-D-Arg-NH2 reduced WNV infection in primary neurons. We have also shown that myelin basic protein, a deficiency of which is linked to neurological abnormalities of the brain, is sensitive to NS3 proteolysis in vitro and therefore this protein represents a convenient test substrate for the studies of NS3. A three-dimensional model of WNV NS3 that we created may provide a structural guidance and a rationale for the subsequent design of fine-tuned inhibitors. Overall, our findings represent a foundation for in-depth mechanistic and structural studies as well as for the design of novel and efficient inhibitors of WNV NS3.
Shiryaev
Sergey A
SA
The Burnham Institute, La Jolla, CA 92037, USA.
Ratnikov
Boris I
BI
Chekanov
Alexei V
AV
Sikora
Sergey
S
Rozanov
Dmitri V
DV
Godzik
Adam
A
Wang
Jun
J
Smith
Jeffrey W
JW
Huang
Ziwei
Z
Lindberg
Iris
I
Samuel
Melanie A
MA
Diamond
Michael S
MS
Strongin
Alex Y
AY
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000032:analysis* / Q000737:chemistry; D002478:Cells, Cultured; D017124:Conserved Sequence; D045683:Furin / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009186:Myelin Sheath / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D011480:Protease Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D020365:RNA Helicases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015843:Serpins / Q000494:pharmacology; D013379:Substrate Specificity; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014902:West Nile virus / Q000201:enzymology* / Q000235:genetics; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D001120:Arginine; D045683:Furin; D010455:Peptides; D011480:Protease Inhibitors; D020365:RNA Helicases; D012697:Serine Endopeptidases; D015843:Serpins; D017361:Viral Nonstructural Proteins; C060028:NS3 protein, flavivirus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases