Identification of a novel peptide substrate of HSV-1 protease using substrate phage display.
The method of substrate phage display was used to select a preferred substrate from three monovalent display libraries using the HSV-1 protease. The display libraries consisted of four random amino acids, six random amino acids, and a biased library containing four amino acids from the P side of the HSV-1 maturation site followed by four random amino acids. A series of consensus peptides was synthesized based upon the results from these screens and tested in peptide cleavage assays. An eight amino acids consensus peptide (LVLASSSF) derived from the phage results was cleaved as efficiently as a 20-mer maturation site peptide. The selected amino acid sequences also allowed the design of a four amino acid paranitroanilide substrate for continuous assay of HSV-1 protease. Similar to HCMV protease, these results define P4 to P1 as a minimal substrate recognition domain for the HSV-1 protease and suggest that P4 to P1 is the minimal substrate domain which all herpesvirus proteases recognize.
O'Boyle
D R
DR
Bristol-Myers Squibb Pharmaceutical Research Institute, Wallingford, Connecticut 06492, USA.
Pokornowski
K A
KA
McCann
P J
PJ
Weinheimer
S P
SP
D000595:Amino Acid Sequence; D017104:Bacteriophage M13 / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D002213:Capsid / Q000235:genetics / Q000378:metabolism*; D016384:Consensus Sequence; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D018259:Herpesvirus 1, Human / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism; D019151:Peptide Library; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D014764:Viral Proteins
D017931:DNA Primers; D009842:Oligopeptides; D019151:Peptide Library; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C079287:scaffold protein, Herpes simplex virus-1; D012697:Serine Endopeptidases