Searching sequence space for high-affinity binding peptides using ribosome display.
We present the construction of a synthetic library based on the scaffold of bovine heart fatty acid-binding protein (FABP) with 1.1x10(14) independent members. Ribosome display was applied to select streptavidin-binding peptides in vitro from 2x10(13) molecules of the library each encoding FABP with 15 contiguous random amino acid residues at its N terminus. The selection yielded several different binding peptides. The best binder possessed a dissociation constant as low as 4nM and, in contrast to the previously isolated peptides, contained no HPQ motif. A substitution analysis enabled shortening of the 15-mer peptide and revealed a 9-mer variant with a dissociation constant of 17nM, which is a 1000-fold increase of affinity compared to the already known peptides of this size. This high-affinity binding peptide in combination with the whole set of streptavidin conjugates should be an extremely useful tool for the detection and purification of recombinant proteins.
Lamla
Thorsten
T
Institut für Biochemie, Freie Universität Berlin, Germany.
Erdmann
Volker A
VA
D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D002417:Cattle; D017931:DNA Primers / Q000737:chemistry; D050556:Fatty Acid-Binding Proteins; D006321:Heart / Q000502:physiology*; D066298:In Vitro Techniques; D009363:Neoplasm Proteins; D009842:Oligopeptides / Q000737:chemistry*; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism*; D019809:Streptavidin / Q000378:metabolism*; D014158:Transcription, Genetic
D002352:Carrier Proteins; D017931:DNA Primers; D050556:Fatty Acid-Binding Proteins; D009363:Neoplasm Proteins; D009842:Oligopeptides; D019151:Peptide Library; D010455:Peptides; D012333:RNA, Messenger; D019809:Streptavidin