ABSTRACT
A random hexapeptide library (one-bead-one-compound), containing sixteen amino acids (16(6) different sequences) was synthesized on a Tentagel resin previously modified with a dipeptide linker (Asp-Pro). This peptide bond is highly susceptible to cleavage under mild acidic conditions in a salt-free solution prepared with H(2)(16)O/H(2)(18)O (60/40% v/v). In the hydrolysis, hexapeptides are released with an additional Asp residue partially labeled with (18)O at the C-terminus. These conditions are fully compatible with ESI-MS analysis and facilitate sequencing by MS, as N- and C-terminal ions can be easily differentiated in MS/MS spectra. The peptides were sequenced manually and also with de novo sequencing programs, and identifying them in a database containing all possible heptapeptide sequences or in a filtered database. The proposed strategy is also compatible with stepwise Edman degradation using either intact beads or the released free peptides.