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 1.1 X 10(14) independent members. Ribosome display was applied to select streptavidin-binding peptides in vitro from 2 X 10(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 4 nM 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 17 nM, 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. (C) 2003 Elsevier Science Ltd. All rights reserved.