PURIFICATION OF SYNTHETIC PEPTIDE LIBRARIES BY AFFINITY-CHROMATOGRAPHY USING THE AVIDIN-BIOTIN SYSTEM

The specific interaction between biotin and avidin was exploited in th e affinity purification of solid-phase synthesized peptide libraries. During peptide library synthesis, by means of the single-resin method in which coupling on variable positions is carried out using an equimo lar mixture of amino acids, biotin was used to cap the unreacted amino groups remaining after coupling of the equimolar amino acid mixture. The following synthesis and deprotection procedures were performed as usual in tert,-butyloxycarbonyl chemistry. The purification of the pep tide mixture containing N-biotinylated sequences was performed by affi nity chromatography on an avidin-agarose column. The unwanted terminat ed sequences were retained in the avidin column while the purified pep tide mixture was eluted as indicated by reverse-phase HPLC and MS anal ysis monitoring. The avidin column was regenerated and the biotinylate d sequences were released under reversible denaturing conditions. The affinity column was then reusable for repetitive purifications. The us efulness of biotinylation for peptide library purification is demonstr ated here for the first time for a peptide mixture containing by-produ cts that cannot be separated from the mixture by classical HPLC purifi cation. This purification technique could be applied to all syntheses, presenting difficult reacting steps. (C) 1995 Academic Press, Inc.