UNEXPECTED FRAMESHIFTS FROM GENE TO EXPRESSED PROTEIN IN A PHAGE-DISPLAYED PEPTIDE LIBRARY

A library of long peptides displayed on the pm protein of filamentous phage was used in biopanning experiments against several protein targe ts. We find that a large percentage of phage clones that bind specific ally to a target contain peptide-encoding genes that do not have an OR F. Instead, the reading frame is either interrupted by one or more non suppressed stop codons, or a post-transcriptional frameshift is needed to account for the expression of the minor phage coat protein pIII. T he percentage of frameshifted clones varies depending on the target, I t can be as high as 90% for clones specific for soluble forms of certa in cytokine receptors. Conversely, biopanning against four mAbs did no t yield any frameshifted clones. Our studies focused on one clone that binds specifically to rat growth hormone binding protein (GHBP) yet d oes not have an ORF. ih secondary peptide library containing random mu tations of this sequence was constructed and panned against GHBP to op timize and correct the reading frame. In the last round (roland two) o f panning with this library, none of the phage clones that bound to GH BP had an ORF. However, careful analysis of these clones allowed us to design a synthetic peptide capable of binding to GHBP. The results of this study indicate that ORFs are not required to obtain gene express ion of the minor coat protein of filamentous phage and suggest that so me ORF- clones may have a selective advantage over the clones having O RFs.