Combinatorial Cellulose-Bound Peptide Libraries: Screening Tools for the Identification of Peptides That Bind Ligands with Predefined Specificity

We describe the synthesis of structurally different types of combinatorial peptide libraries on continuous cellulose membrane supports. These libraries consisting of tens of millions of different peptides were screened for their ability to bind given ligands such as the monoclonal antibody Tab2, transforming growth factor-β (TGFβ), nickel(II) (Ni²⁺), and technetium-99m (^99mTc). We were thus able to detect the linear transforming growth factor-α (TFGα) epitope SHFND recognized by Tab2. Other peptides that also bound Tab2 were identified within the same experiment. A first screening step for identification of peptide mixtures that bind to other ligands of biological interest is shown for peptide mixtures XB₁XB₂XX (B = defined amino acid, X = randomized position) that bind to Ni²⁺ and ^99mTc. A combinatorial linear all L- or all D-library XXB₁B₂XX and two libraries conformationally restrained either via a disulfide bridge between a C-and an N-terminal cysteine cyclo(C¹-C⁸)-C¹XXB₁B₂XXC⁸] or an amide bond between the alpha amino group of the N-terminus and the gamma carboxyl group of a C-terminal glutamic acid cyclo(X¹-E⁷)-X¹XB₁B₂XXE⁷] were screened with transforming growth factor-β, resulting in structurally different peptides mixtures that bound to this ligand. The results obtained indicate that chemically different types of cellulose-bound combinatorial libraries can be prepared easily, allowing the rapid and inexpensive screening of millions of peptides for selection of single molecules with predefined specificity that bind to given ligands such as proteins, metals, nucleic acids, and other molecules of biological interest.