Inhibition of the bovine viral diarrhoea virus NS3 serine protease by a boron-modified peptidyl mimetic of its natural substrate.
Bovine viral diarrhoea virus (BVDV) is closely related to hepatitis C virus (HCV), and has been used as a surrogate virus in drug development for HCV infection. Similar to HCV, BVDV-encoded NS3 serine proteinase is responsible for multiple cleavages in the viral polyprotein, generating mature NS4A, NS4B, NS5A and NS5B proteins. NS3-dependent cleavage sites of BVDV contain a strictly conserved leucine at P1, and either serine or alanine at P1'. The full length BVDV NS3/4A serine protease has been cloned and expressed in bacterial cells. The enzyme has been purified from the soluble portion of Escherichia coli via a two-step purification procedure employing chromatography on heparin resin and gel filtration. The protease activity was characterized using in vitro translated BVDV NS4A/B and NS5A/B polyprotein substrates. A boronic acid analogue of the BVDV NS4A/NS4B cleavage site was synthesized and shown to be an efficient inhibitor of the NS3 serine protease in vitro. The compound, designated DPC-AB9144-00, inhibited approximately 75% of the NS3/4 activity at 10 microM with the NS4A/B substrate. However, no antiviral activity was detected with DPC-AB9144-00 in BVDV-infected Madin-Darby bovine kidney cells at concentrations as great as 90 pM, suggesting permeability or that other cellular-derived limitations were present.
Bukhtiyarova
M
M
The Experimental Station, Bristol-Myers Squibb, Wilmington, Del, USA.
Rizzo
C J
CJ
Kettner
C A
CA
Korant
B D
BD
Scarnati
H T
HT
King
R W
RW
D000818:Animals; D001667:Binding, Competitive; D015153:Blotting, Western; D001895:Boron / Q000737:chemistry*; D002417:Cattle; D002460:Cell Line; D001908:Diarrhea Viruses, Bovine Viral / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D018716:Molecular Mimicry; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D012697:Serine Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000737:chemistry* / Q000494:pharmacology*; D014779:Virus Replication / Q000187:drug effects
D010455:Peptides; D015842:Serine Proteinase Inhibitors; D012697:Serine Endopeptidases; D001895:Boron