Method of recovering a nucleic acid encoding a proteinaceous binding domain which binds a target material

A method of recovering a nucleic acid encoding a binding domain, the method comprising:

• providing a variegated population of filamentous phage, wherein each phage displays on its outer surface a potential binding domain that comprises an antibody domain and is physically associated with a nucleic acid sequence that encodes the potential binding domain, the encoded potential binding domains differ from one another through the at least partially random variation of one or more amino acids corresponding to a hypervariable region, and the random variation of at least one of the amino acid positions is by random selection of the codon encoding the amino acid at said position from a set of codons, the set being characterized by one or more of the following properties:
• (a) the set includes at least one codon for each of at least two different amino acids other than cysteine, and excludes all codons encoding cysteine,
• (b) the set provides a single codon for each encoded amino acid, and
• (c) the amino acids encoded by the set are represented at substantially equal frequency;
• contacting the phage with a target material such that the potential binding domains and the target material may interact while each potential binding domain and a nucleic acid sequence that encodes it remain physically associated;
• isolating a binding domain that binds to the target material;
• and
• recovering the particular nucleic acid that is physically associated with the isolated binding domain during the contacting.

A process for determining a binding property of a proteinaceous domain, the process comprising:

• mutagenizing a gene encoding an antibody domain to form a gene encoding a potential binding domain, wherein at least one codon encoding an amino acid in a hypervariable region is mutagenized;
• displaying the potential binding domain on the outer surface of an amplifiable genetic package, wherein said amplifiable genetic package is a filamentous bacteriophage that contains the gene encoding said potential binding domain,
• contacting the package with the target material, and
• determining whether the package displaying the potential binding domain binds to said target material.

A method of isolating a binding protein, the method comprising:

• preparing a variegated population of filamentous phage, each phage including a nucleic acid construct that encodes a chimeric protein that comprises a potential binding domain and a polypeptide encoded by a filamentous phage gene VIII or a filamentous phage gene III, the encoded potential binding domains differ through the at least partially random variation of one or more amino acid positions of a parental binding domain;
• expressing the potential binding domains encoded by the nucleic acid constructs of the phage population;
• isolating at least one phage from the population, the isolated phage including a nucleic acid construct that encodes a potential binding domain that binds to the target material with at least a predetermined affinity;
• and
• recovering the nucleic acid construct from the isolated phage.