De novo proteins from binary-patterned combinatorial libraries.

Combinatorial libraries of well-folded de novo proteins can provide a rich source of reagents for the isolation of novel molecules for biotechnology and medicine. To produce libraries containing an abundance of well-folded sequences, we have developed a method that incorporates both rational design and combinatorial diversity. Our method specifies the "binary patterning" of polar and nonpolar amino acids, but allows combinatorial diversity of amino acid side chains at each polar and nonpolar site in the sequence. Protein design by binary patterning is based on the premise that the appropriate arrangement of polar and nonpolar residues can direct a polypeptide chain to fold into amphipathic elements of secondary structures, which anneal together to form a desired tertiary structure. A designed binary pattern exploits the periodicities inherent in protein secondary structure, while allowing the identity of the side chain at each polar and non-polar position to be varied combinatorially. This chapter provides an overview of the considerations necessary to design binary patterned libraries of novel proteins.