GENERATION AND SCREENING OF COMBINATORIAL PEPTIDE LIBRARIES DESIGNED FOR RAPID SEQUENCING BY MASS-SPECTROMETRY

We have developed a method for rapidly sequencing compounds isolated from support-bound combinatorial libraries. During the synthesis of the library, a capping reagent is used to effect partial termination at each coupling step. In this way, each of the resin beads carries not only the full-length product but also small amounts of sequence-specific termination products. The sequence of the full-length product is determined with matrix-assisted laser desorption ionization mass spectrometry, which is used to measure mass differences between adjacent members of the termination series. The observed mass differences identify the corresponding monomers. With mixtures of terminating reagents, monomers of identical mass can also be distinguished. This method is fast, accurate, and reliable, and it is compatible with a wide range of unnatural synthetic libraries. Because our sequencing method directly analyzes the compounds used for screening, synthetic byproducts can be easily identified and, with this information, reaction conditions can be optimized to minimize them. To demonstrate the utility of the method, we have prepared combinatorial libraries of acetylated and non-acetylated pentapeptides on polystyrene beads. These libraries were used to identify ligands to an HTV-neutralizing antibody and to streptavidin.