A novel combinatorial strategy for the redesign of proteins based on the strength and specificity of intra- and interprotein interactions is described. The strategy has been used to redesign the hydrophobic core of the B domain of protein A. Using one-bead-one-compound combinatorial chemistry, 300 analogues of the C-terminal a-helix of the B domain, H3(x), have been synthesized using a biocompatible resin and the HMFS linker, allowing the screening to occur while the peptides were bound to the resin. The screening was based on the ability of the H3(x) analogues to interact with the N-terminal helices of the B domain, H1-H2, and retain the native B domain activity, that is binding to IgG. Eight different analogues containing some nonconservative mutations were obtained from the library, the two most frequent of which, H3(P1) and H3(P2), were studied in detail. CD analysis revealed that the active analogues interact with H1-H2. To validate the redesign strategy the covalent modified domains H1-H2-H3(P1) and H1-H2-H3(P2) were synthesized and characterized. CD and NMR analysis revealed that they had a unique, stable, and well-defined three-dimensional structure similar to that for the wild-type B domain, This combinatorial strategy allows us to select for redesigned proteins with the desired activity or the desired physicochemical properties provided the right screening test is used. Furthermore, it is rich in potential for the chemical modification of proteins overcoming the drawbacks associated with the total synthesis of large protein domains.