title: Detection of small-molecule enzyme inhibitors with peptides isolated from phage-displayed combinatorial peptide libraries.
abstract: BACKGROUND
  The rapidly expanding list of pharmacologically important targets has highlighted the need for ways to discover new inhibitors that are independent of functional assays. We have utilized peptides to detect inhibitors of protein function. We hypothesized that most peptide ligands identified by phage display would bind to regions of biological interaction in target proteins and that these peptides could be used as sensitive probes for detecting low molecular weight inhibitors that bind to these sites.
  
  
  RESULTS
  We selected a broad range of enzymes as targets for phage display and isolated a series of peptides that bound specifically to each target. Peptide ligands for each target contained similar amino acid sequences and competition analysis indicated that they bound one or two sites per target. Of 17 peptides tested, 13 were found to be specific inhibitors of enzyme function. Finally, we used two peptides specific for Haemophilus influenzae tyrosyl-tRNA synthetase to show that a simple binding assay can be used to detect small-molecule inhibitors with potencies in the micromolar to nanomolar range.
  
  
  CONCLUSIONS
  Peptidic surrogate ligands identified using phage display are preferentially targeted to a limited number of sites that inhibit enzyme function. These peptides can be utilized in a binding assay as a rapid and sensitive method to detect small-molecule inhibitors of target protein function. The binding assay can be used with a variety of detection systems and is readily adaptable to automation, making this platform ideal for high-throughput screening of compound libraries for drug discovery.
authors.affiliation: Novalon Pharmaceutical Corporation, Durham, NC 27703, USA.
authors.forename: R
authors.initials: R
authors.lastname: Hyde-DeRuyscher
authors.forename: L A
authors.initials: LA
authors.lastname: Paige
authors.forename: D J
authors.initials: DJ
authors.lastname: Christensen
authors.forename: N
authors.initials: N
authors.lastname: Hyde-DeRuyscher
authors.forename: A
authors.initials: A
authors.lastname: Lim
authors.forename: Z L
authors.initials: ZL
authors.lastname: Fredericks
authors.forename: J
authors.initials: J
authors.lastname: Kranz
authors.forename: P
authors.initials: P
authors.lastname: Gallant
authors.forename: J
authors.initials: J
authors.lastname: Zhang
authors.forename: S M
authors.initials: SM
authors.lastname: Rocklage
authors.forename: D M
authors.initials: DM
authors.lastname: Fowlkes
authors.forename: P A
authors.initials: PA
authors.lastname: Wendler
authors.forename: P T
authors.initials: PT
authors.lastname: Hamilton
mesh_terms: D000426:Alcohol Dehydrogenase / Q000037:antagonists & inhibitors; D000595:Amino Acid Sequence; D000604:Amino Acyl-tRNA Synthetases / Q000037:antagonists & inhibitors; D001435:Bacteriophages / Q000378:metabolism*; D001710:Biotin / Q000737:chemistry; D043424:Carboxypeptidase B; D002268:Carboxypeptidases / Q000037:antagonists & inhibitors; D002846:Chromatography, Affinity; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000302:isolation & purification / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006193:Haemophilus influenzae / Q000201:enzymology; D006593:Hexokinase / Q000037:antagonists & inhibitors; D007700:Kinetics; D008024:Ligands; D008969:Molecular Sequence Data; D019151:Peptide Library; D006005:Phosphorylases / Q000037:antagonists & inhibitors; D014448:Tyrosine-tRNA Ligase / Q000037:antagonists & inhibitors; D001617:beta-Glucosidase / Q000037:antagonists & inhibitors
chemical_list: D004791:Enzyme Inhibitors; D008024:Ligands; D019151:Peptide Library; D001710:Biotin; D000426:Alcohol Dehydrogenase; D006005:Phosphorylases; D006593:Hexokinase; D001617:beta-Glucosidase; D002268:Carboxypeptidases; D043424:Carboxypeptidase B; D000604:Amino Acyl-tRNA Synthetases; D014448:Tyrosine-tRNA Ligase; C100134:prolyl T RNA synthetase