Redesigning the substrate specificity of the hepatitis C virus NS3 protease.
BACKGROUND
Hepatitis C Virus (HCV) non-structural protein 3 (NS3) encodes a trypsin-like serine protease that catalyzes the cleavages at the NS3/NS4A, NS4A/NS4B, NS4B/NS5A and NS5A/NS5B junctions in the viral polyprotein and that shows a preference for a cysteine as the P1 residue.


RESULTS
We describe here a partial model of the HCV NS3 protease which allowed us to predict the position of the secondary structure elements of the enzyme and of the residues involved in its specificity. By replacing these with the corresponding residues of Streptomyces griseus protease B1 we obtained a protease that, similar to the bacterial protein and unlike the wild-type enzyme, is able to cleave a substrate containing a phenylalanine in the P1 position.


CONCLUSIONS
These results confirm the reliability of our model and represent one of the few examples of redesign of a serine protease substrate specificity directed by molecular modelling.
Failla
C M
CM
Istituto di Ricerche di Biologia Molecolare (IRBM) P. Angeletti, Pomezia, Italy.
Pizzi
E
E
De Francesco
R
R
Tramontano
A
A
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D015195:Drug Design; D006367:HeLa Cells; D016174:Hepacivirus / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013305:Streptomyces griseus / Q000201:enzymology / Q000235:genetics; D013379:Substrate Specificity; D017361:Viral Nonstructural Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases